Methods and compositions for modulating a genome

ABSTRACT

Methods and compositions for modulating a target genome are disclosed.

This application is a Continuation of International Application No.PCT/US2019/048607, filed Aug. 28, 2019, which claims priority to U.S.Ser. No. 62/723,886 filed Aug. 28, 2018, U.S. Ser. No. 62/725,778 filedAug. 31, 2018, U.S. Ser. No. 62/850,883 filed May 21, 2019, and U.S.Ser. No. 62/864,924 filed Jun. 21, 2019, the entire contents of each ofwhich is incorporated herein by reference.

SEQUENCE LISTING

The instant application contains a Sequence Listing which has beensubmitted electronically in ASCII format and is hereby incorporated byreference in its entirety. Said ASCII copy, created on Aug. 28, 2019, isnamed V2065-7000WO_SL.txt and is 4,004,548 bytes in size.

BACKGROUND

Integration of a nucleic acid of interest into a genome occurs at lowfrequency and with little site specificity, in the absence of aspecialized protein to promote the insertion event. Some existingapproaches, like CRISPR/Cas9, are more suited for small edits and areless effective at integrating longer sequences. Other existingapproaches, like Cre/loxP, require a first step of inserting a loxP siteinto the genome and then a second step of inserting a sequence ofinterest into the loxP site. There is a need in the art for improvedproteins for inserting sequences of interest into a genome.

SUMMARY OF THE INVENTION

This disclosure relates to novel compositions, systems and methods foraltering a genome at one or more locations in a host cell, tissue orsubject, in vivo or in vitro. In particular, the invention featurescompositions, systems and methods for the introduction of exogenousgenetic elements into a host genome.

Features of the compositions or methods can include one or more of thefollowing enumerated embodiments.

1. A system for modifying DNA comprising:

(a) a polypeptide or a nucleic acid encoding a polypeptide, wherein thepolypeptide comprises (i) a reverse transcriptase domain and (ii) anendonuclease domain; and

(b) a template RNA (or DNA encoding the template RNA) comprising (i) asequence that binds the polypeptide and (ii) a heterologous objectsequence that encodes a therapeutic polypeptide or that encodes amammalian (e.g., human) polypeptide, or a fragment or variant

thereof.2. A system for modifying DNA comprising:

(a) a polypeptide or a nucleic acid encoding a polypeptide, wherein thepolypeptide comprises (i) a reverse transcriptase domain and (ii) anendonuclease domain; and

(b) a template RNA (or DNA encoding the template RNA) comprising (i) asequence that binds the polypeptide and (ii) a heterologous objectsequence, wherein one or more of:

-   -   i. the heterologous object sequence encodes a protein, e.g. an        enzyme (e.g., a lysosomal enzyme) or a blood factor (e.g.,        Factor I, II, V, VII, X, XI, XII or XIII);    -   ii. the heterologous object sequence comprises a tissue specific        promoter or enhancer;    -   iii. the heterologous object sequence encodes a polypeptide of        greater than 250, 300, 400, 500, or 1,000 amino acids, and        optionally up to 7,500 amino acids;    -   iv. the heterologous object sequence encodes a fragment of a        mammalian gene but does not encode the full mammalian gene,        e.g., encodes one or more exons but does not encode a        full-length protein;    -   v. the heterologous object sequence encodes one or more introns;    -   vi. the heterologous object sequence is other than a GFP, e.g.,        is other than a fluorescent protein or is other than a reporter        protein.    -   vii. the heterologous object sequence is other than a T cell        chimeric antigen receptor        3. A system for modifying DNA comprising:

(a) a polypeptide or a nucleic acid encoding a polypeptide, wherein thepolypeptide comprises (i) a reverse transcriptase domain and (ii) anendonuclease domain; and

(b) a template RNA (or DNA encoding the template RNA) comprising (i) asequence that binds the polypeptide and (ii) a heterologous objectsequence.

4. A system for modifying DNA comprising:

(a) a polypeptide or a nucleic acid encoding a polypeptide, wherein thepolypeptide comprises (i) a target DNA binding domain, (ii) a reversetranscriptase domain and (iii) an endonuclease domain; and

(b) a template RNA (or DNA encoding the template RNA) comprising (i) asequence that binds the polypeptide and (ii) a heterologous objectsequence.

5. A system for modifying DNA comprising:

(a) a polypeptide or a nucleic acid encoding a polypeptide, wherein thepolypeptide comprises (i) a reverse transcriptase domain and (ii) anendonuclease domain, wherein one or both of (i) or (ii) are derived froman avian retrotransposase, e.g., have a sequence of Table 2 or 3 or atleast 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identitythereto; and

(b) a template RNA (or DNA encoding the template RNA) comprising (i) asequence that binds the polypeptide and (ii) a heterologous objectsequence.

6. A system for modifying DNA comprising:

(a) a polypeptide or a nucleic acid encoding a polypeptide, wherein thepolypeptide comprises (i) a reverse transcriptase domain and (ii) anendonuclease domain, wherein the polypeptide has an activity at 37° C.that is no less than 70%, 75%, 80%, 85%, 90%, or 95% of its activity at25° C. under otherwise similar conditions; and

(b) a template RNA (or DNA encoding the template RNA) comprising (i) asequence that binds the polypeptide and (ii) a heterologous objectsequence.

7. The system of embodiment 6, wherein the polypeptide is derived froman avian retrotransposase, e.g., an avian retrotransposase of column 8of Table 3, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%,96%, 97%, 98%, or 99% identity thereto.8. The system of embodiment 6, wherein the avian retrotransposase is aretrotransposase from Taeniopygia guttata, Geospiza fortis, Zonotrichiaalbicollis, or Tinamus guttatus, or a sequence having at least 70%, 75%,80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.9. The system of embodiment 6, wherein the polypeptide is derived from aretrotransposase of column 8 of Table 3, or a sequence having at least70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.10. The system of any of the preceding embodiments, wherein the templateRNA comprises a sequence of Table 3 (e.g., one or both of a 5′untranslated region of column 6 of Table 3 and a 3′ untranslated regionof column 7 of Table 3), or a sequence having at least 70%, 75%, 80%,85%, 90%, 95%, 96%, 97%, 98%, or 99% identity thereto.11. A system for modifying DNA comprising:

(a) a polypeptide or a nucleic acid encoding a polypeptide, wherein thepolypeptide comprises (i) a reverse transcriptase domain and (ii) anendonuclease domain; and

(b) a template RNA (or DNA encoding the template RNA) comprising (i) asequence that binds the polypeptide and (ii) a heterologous objectsequence, wherein one or more of:

-   -   i. the nucleic acid encoding the polypeptide and the template        RNA or a nucleic acid encoding the template RNA are separate        nucleic acids;    -   ii. the template RNA does not encode an active reverse        transcriptase, e.g., comprises an inactivated mutant reverse        transcriptase, e.g., as described in Examples 1-2, or does not        comprise a reverse transcriptase sequence; or    -   iii. the template RNA does not encode an active endonuclease,        e.g., comprises an inactivated endonuclease or does not comprise        an endonuclease; or    -   iv. the template RNA comprises one or more chemical        modifications.        12. A system for modifying DNA comprising:

(a) a polypeptide or a nucleic acid encoding a polypeptide, wherein thepolypeptide comprises (i) a reverse transcriptase domain and (ii) anendonuclease domain; and

(b) a template RNA (or DNA encoding the template RNA) comprising (i) a5′ untranslated sequence that binds the polypeptide, (ii) a 3′untranslated sequence that binds the polypeptide, (iii) a heterologousobject sequence, and (iv) a promoter operably linked to the heterologousobject sequence,

wherein the promoter is disposed between the 5′ untranslated sequencethat binds the polypeptide and the heterologous sequence, or

wherein the promoter is disposed between the 3′ untranslated sequencethat binds the polypeptide and the heterologous sequence.

13. A system for modifying DNA comprising:

(a) a polypeptide or a nucleic acid encoding a polypeptide, wherein thepolypeptide comprises (i) a reverse transcriptase domain and (ii) anendonuclease domain; and

(b) a template RNA (or DNA encoding the template RNA) comprising (i) a5′ untranslated sequence that binds the polypeptide, (ii) a 3′untranslated sequence that binds the polypeptide, and (iii) aheterologous object sequence, and

wherein the heterologous object sequence comprises an open reading frame(or the reverse complement thereof) in a 5′ to 3′ orientation on thetemplate RNA; or

wherein the heterologous object sequence comprises an open reading frame(or the reverse complement thereof) in a 3′ to 5′ orientation on thetemplate RNA.

14. A system for modifying DNA comprising:

(a) a polypeptide or a nucleic acid encoding a polypeptide, wherein thepolypeptide comprises (i) a reverse transcriptase domain and (ii) anendonuclease domain, wherein at least one of (i) or (ii) isheterologous, and

(b) a template RNA (or DNA encoding the template RNA) comprising (i) asequence that binds the polypeptide and (ii) a heterologous objectsequence.

15. A system for modifying DNA comprising:

(a) a polypeptide or a nucleic acid encoding a polypeptide, wherein thepolypeptide comprises (i) a target DNA binding domain, (i) a reversetranscriptase domain and (iii) an endonuclease domain, wherein at leastone of (i), (ii) or (iii) is heterologous, and

(b) a template RNA (or DNA encoding the template RNA) comprising (i) asequence that binds the polypeptide and (ii) a heterologous objectsequence.

16. A system for modifying DNA comprising:

(a) a polypeptide or a nucleic acid encoding a polypeptide, wherein thepolypeptide comprises (i) a sequence at least 80% identical (e.g., atleast 85%, 90%, 95%, 97%, 98%, 99%, 100% identical) to a reversetranscriptase domain of a purinic/apyrimidinic endonuclease (APE)-typenon-LTR retrotransposon and (ii) a sequence at least 80% identical(e.g., at least 85%, 90%, 95%, 97%, 98%, 99%, 100% identical) to anendonuclease domain of an APE-type non-LTR retrotransposon; and

(b) a template RNA (or DNA encoding the template RNA) comprising (i) asequence that binds the polypeptide and (ii) a heterologous objectsequence.

17. A system for modifying DNA comprising:

(a) a polypeptide or a nucleic acid encoding a polypeptide, wherein thepolypeptide comprises (i) a sequence at least 80% identical (e.g., atleast 85%, 90%, 95%, 97%, 98%, 99%, 100% identical) to a reversetranscriptase domain of a restriction enzyme-like endonuclease(RLE)-type non-LTR retrotransposon, (ii) a sequence at least 80%identical (e.g., at least 85%, 90%, 95%, 97%, 98%, 99%, 100% identical)to an endonuclease domain of a RLE-type non-LTR retrotransposon, and(iii) a heterologous target DNA binding domain (e.g., a heterologouszinc-finger DNA binding domain); and

(b) a template RNA (or DNA encoding the template RNA) comprising (i) asequence that binds the polypeptide and (ii) a heterologous objectsequence.

18. The system of any of the preceding embodiments, wherein the templateRNA comprises (iii) a promoter operably linked to the heterologousobject sequence.19. The system of any of the preceding embodiments, wherein thepolypeptide further comprises (iii) a DNA-binding domain.20. The system of embodiment 17, wherein the polypeptide comprises asequence at least 80% identical (e.g., at least 85%, 90%, 95%, 97%, 98%,99%, 100% identical) to the sequence of SEQ ID NO: 1016.21. The system of any of the preceding embodiments, wherein thepolypeptide comprises a sequence at least 80% identical (e.g., at least85%, 90%, 95%, 97%, 98%, 99%, 100% identical) to a sequence in column 8of Table 3.22. The system of any of the preceding embodiments, wherein the nucleicacid encoding the polypeptide and the template RNA or the nucleic acidencoding the template RNA are covalently linked, e.g., are part of afusion nucleic acid.23. The system of embodiment 22, wherein the fusion nucleic acidcomprises RNA.24. The system of embodiment 22, wherein the fusion nucleic acidcomprises DNA.25. The system of any of the preceding embodiments, wherein (b)comprises template RNA.26. The system of embodiment 25, wherein the template RNA furthercomprises a nuclear localization signal.27. The system of any of the preceding embodiments, wherein (a)comprises RNA encoding the polypeptide.28. The system of embodiment 27, wherein the RNA of (a) and the RNA of(b) are separate RNA molecules.29. The system of embodiment 28, wherein the RNA of (a) and the RNA of(b) are present at a ratio of between 10:1 and 5:1, 5:1 and 2:1, 2:1 and1:1, 1:1 and 1:2, 1:2 and 1:5, or 1:5 and 1:10.30. The system of embodiment 28, wherein the RNA of (a) does notcomprise a nuclear localization signal.31. The system of any of the preceding embodiments, wherein thepolypeptide further comprises a nuclear localization signal and/or anucleolar localization signal.32. The system of any of the preceding embodiments, wherein (a)comprises an RNA that encodes: (i) the polypeptide and (ii) a nuclearlocalization signal and/or a nucleolar localization signal.33. The system of any of the preceding embodiments, wherein the RNAcomprises a pseudoknot sequence, e.g., 5′ of the heterologous objectsequence.34. The system of embodiment 33, wherein the RNA comprises a stem-loopsequence or a helix, 5′ of the pseudoknot sequence.35. The system of embodiment 33 or 34, wherein the RNA comprises one ormore (e.g., 2, 3, or more) stem-loop sequences or helices 3′ of thepseudoknot sequence, e.g. 3′ of the pseudoknot sequence and 5′ of theheterologous object sequence.36. The system of any of embodiments 33-35, wherein the template RNAcomprising the pseudoknot has catalytic activity, e.g., RNA-cleavingactivity, e.g, cis-RNA-cleaving activity.37. The system of any of the preceding embodiments, wherein the RNAcomprises at least one stem-loop sequence or helix, e.g., 3′ of theheterologous object sequence, e.g. 1, 2, 3, 4, 5 or more stem-loopsequences, hairpins or helices sequences.38. Any above-numbered system, wherein the polypeptide comprises asequence of at least 50 amino acids (e.g., at least 100, 150, 200, 300,500 amino acids) having at least 80% identity (e.g., at least 85%, 90%,95%, 97%, 98%, 99%, 100% identity) to a sequence of a polypeptide listedin Table 1, or a reverse transcriptase domain or endonuclease domainthereof.39. Any above-numbered system, wherein the polypeptide comprises asequence of at least 50 amino acids (e.g., at least 100, 150, 200, 300,500 amino acids) having at least 80% identity (e.g., at least 85%, 90%,95%, 97%, 98%, 99%, 100% identity) to a sequence of a polypeptide listedin any of Tables 2-3 or a reverse transcriptase domain, endonucleasedomain, or DNA binding domain thereof.40. Any above-numbered system, wherein the polypeptide comprises asequence of at least 50 amino acids (e.g., at least 100, 150, 200, 300,500 amino acids) having at least 80% identity (e.g., at least 85%, 90%,95%, 97%, 98%, 99%, 100% identity) to the amino acid sequence of column8 of Table 3, or a reverse transcriptase domain, endonuclease domain, orDNA binding domain thereof.41. Any above-numbered system, wherein the template RNA comprises asequence of Table 3 (e.g., one or both of a 5′ untranslated region ofcolumn 6 of Table 3 and a 3′ untranslated region of column 7 of Table3), or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%,97%, 98%, or 99% identity thereto.42. The system of embodiment 41, wherein the template RNA comprises asequence of about 100-125 bp from a 3′ untranslated region of column 7of Table 3, e.g., wherein the sequence comprises nucleotides 1-100,101-200, or 201-325 of the 3′ untranslated region of column 7 of Table3, or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%,98%, or 99% identity thereto.43. Any above-numbered system, wherein (a) comprises RNA and (b)comprises RNA.44. Any above-numbered system, which comprises only RNA, or whichcomprises more RNA than DNA by an RNA:DNA ratio of at least 10:1, 20:1,30:1, 40:1, 50:1, 60:1, 70:1, 80:1, 90:1, or 100:1.45. Any above-numbered system, which does not comprise DNA, or whichdoes not comprise more than 10%, 5%, 4%, 3%, 2%, or 1% DNA by mass or bymolar amount.46. Any above-numbered system, which is capable of modifying DNA byinsertion of the heterologous object sequence without an interveningDNA-dependent RNA polymerization of (b).47. Any above-numbered system, which is capable of modifying DNA byinsertion of a heterologous object sequence in the presence of aninhibitor of a DNA repair pathway (e.g., SCR7, a PARP inhibitor), or ina cell line deficient for a DNA repair pathway (e.g., a cell linedeficient for the nucleotide excision repair pathway or thehomology-directed repair pathway).48. Any above-numbered system, which does not cause formation of adetectable level of double stranded breaks in a target cell.49. Any above-numbered system, which is capable of modifying DNA usingreverse transcriptase activity, and optionally in the absence ofhomologous recombination activity.50. Any above-numbered system, wherein the template RNA has been treatedto reduce secondary structure, e.g., was heated, e.g., to a temperaturethat reduces secondary structure, e.g., to at least 70, 75, 80, 85, 90,or 95 C.51. The system of embodiment 50, wherein the template RNA wassubsequently cooled, e.g., to a temperature that allows for secondarystructure, e.g, to less than or equal to 30, 25, or 20 C52. A system for modifying DNA comprising:

(a) a polypeptide or a nucleic acid encoding a polypeptide, wherein thepolypeptide comprises (i) a reverse transcriptase domain and (ii) anendonuclease domain; and

(b) a template RNA (or DNA encoding the template RNA) comprising (i) asequence that binds the polypeptide, (ii) a heterologous objectsequence, (iii) a first homology domain having at least 10 bases of 100%identity to a target DNA strand, at the 5′ end of the template RNA, and(iv) a second homology domain having at least 10 bases of 100% identityto a target DNA strand, 5′ end of the template RNA.

53. The system of any of the preceding embodiments, wherein (a) and (b)are part of the same nucleic acid.54. The system of any of embodiments 1-52, wherein (a) and (b) areseparate nucleic acids.55. The system of any of the preceding embodiments, wherein the templateRNA comprises at least 10 bases of 100% identity to a target DNA strand(e.g., wherein the target DNA strand is a human DNA sequence), at the 5′end of the template RNA.56. The system of any of the preceding embodiments, wherein the templateRNA comprises at least 10 bases of 100% identity to a target DNA strand(e.g., wherein the target DNA strand is a human DNA sequence), at the 3′end of the template RNA.57. A host cell (e.g., a mammalian cell, e.g., a human cell) comprisingany preceding numbered system.58. A method of modifying a target DNA strand in a cell, tissue orsubject, comprising administering any preceding numbered system to thecell, tissue or subject, wherein the system reverse transcribes thetemplate RNA sequence into the target DNA strand, thereby modifying thetarget DNA strand.59. The method of embodiment 58, wherein the cell, tissue or subject isa mammalian (e.g., human) cell, tissue or subject.60. The method of any of the preceding embodiments, wherein the cell isa fibroblast.61. The method of any of the preceding embodiments, wherein the cell isa primary cell.62. The method of any of the preceeding embodiments, where in the cellis not immortalized.63. A method of modifying the genome of a mammalian cell, comprisingcontacting the cell with:

(a) a polypeptide or a nucleic acid encoding a polypeptide, wherein thepolypeptide comprises (i) a reverse transcriptase domain, (ii) anendonuclease domain, and optionally (iii) a DNA-binding domain; and

(b) a template RNA (or DNA encoding the template RNA) comprising (i) asequence that binds the polypeptide and (ii) a heterologous objectsequence.

64. The method of embodiment 63, wherein the polypeptide does notcomprise a target DNA binding domain.65. The method of embodiment 63, wherein the polypeptide is derived froman APE-type transposon reverse transcriptase.66. The method of embodiment 63, wherein the (i) a reverse transcriptasedomain (ii) an endonuclease domain, or both of (i) and (ii), have asequence of Table 1 or a sequence having at least 80%, 85%, 90%, 95%,97%, 98%, 99%, 100% identity thereto.67. The method of embodiment 63, wherein the polypeptide furthercomprises a target DNA binding domain.68. A method of modifying the genome of a mammalian cell, comprisingcontacting the cell with:

(a) an RNA encoding a polypeptide, wherein the polypeptide comprises (i)a reverse transcriptase domain, (ii) an endonuclease domain, andoptionally (iii) a DNA-binding domain; and

(b) a template RNA comprising (i) a sequence that binds the polypeptideand (ii) a heterologous object sequence,

wherein the method does not comprise contacting the mammalian cell withDNA, or wherein the compositions of (a) and (b) do not comprise morethan 1%, 0.5%, 0.2%, 0.1%, 0.05%, 0.02%, or 0.01% DNA by mass or bymolar amount of nucleic acid.

69. The method of embodiment 68, which results in the addition of atleast 5, 10, 20, 50, 100, 200, 500, 1,000, 2,000, or 5,000 base pairs ofexogenous DNA sequence to the genome of the mammalian cell.70. The method of embodiment 68 or 69, which results in the addition ofa protein coding sequence to the genome of the mammalian cell.71. A method of inserting DNA into the genome of a mammalian cell,comprising contacting the cell with an RNA composition, wherein the RNAcomposition comprises:

(a) a first RNA that directs insertion of a template RNA into thegenome, and

(b) a template RNA comprising a heterologous sequence, wherein themethod does not comprise contacting the mammalian cell with DNA, or

wherein the compositions of (a) and (b) do not comprise more than 1%,0.5%, 0.2%, 0.1%, 0.05%, 0.02%, or 0.01% DNA by mass or by molar amountof nucleic acid,

wherein the method results in the addition of at least 5, 10, 20, 50,100, 200, 500, 1,000, 2,000, or 5,000 base pairs of DNA (e.g., exogenousDNA) sequence to the genome of the mammalian cell.

72. The method of embodiment 71, wherein the first RNA encodes apolypeptide (e.g., a polypeptide of any of Tables 1, 2, or 3 herein),wherein the polypeptide directs insertion of the template RNA into thegenome.73. The method of embodiments 72, wherein the template RNA furthercomprises a sequence that binds the polypeptide.74. A method of adding at least 5, 10, 15, 20, 25, 30, 35, 40, 45, 50,60, 70, 80, 90, 100, 150, 200, 250, 300, 350, 400, 500, 1000 bp ofexogenous DNA to the genome of a mammalian cell, without delivery of DNAto the cell.75. A method of adding at least 5, 10, 15, 20, 25, 30, 35, 40, 45, 50,60, 70, 80, 90, 100, 150, 200, 250, 300, 350, 400, 500, 1000 bp ofexogenous DNA to the genome of a mammalian cell, wherein the method doesnot comprise contacting the mammalian cell with DNA, or wherein themethod comprises contacting the mammalian cell with a compositioncomprising less than 1%, 0.5%, 0.2%, 0.1%, 0.05%, 0.02%, or 0.01% DNA bymass or by molar amount of nucleic acid.76. A method of adding at least 5, 10, 15, 20, 25, 30, 35, 40, 45, 50,60, 70, 80, 90, 100, 150, 200, 250, 300, 350, 400, 500, 1000 bp ofexogenous DNA to the genome of a mammalian cell, comprising deliveringonly RNA to the mammalian cell.77. A method of adding at least 5, 10, 15, 20, 25, 30, 35, 40, 45, 50,60, 70, 80, 90, 100, 150, 200, 250, 300, 350, 400, 500, 1000 bp ofexogenous DNA to the genome of a mammalian cell, comprising deliveringRNA and protein to the mammalian cell.78. The method of any one of embodiments 68-77, wherein the template RNAserves as the template for insertion of the exogenous DNA.79. The method of any one of embodiments 68-78, which does not compriseDNA-dependent RNA polymerization of exogenous DNA.80. The method of any of embodiments 58-79, which results in theaddition of at least 5, 10, 20, 50, 100, 200, 500, 1,000, 2,000, or5,000 base pairs of DNA to the genome of the mammalian cell.81. The methods of any of embodiments 68-80, wherein the RNA of (a) andthe RNA of (b) are covalently linked, e.g., are part of the sametranscript.82. The methods of any of embodiments 68-80, wherein the RNA of (a) andthe RNA of (b) are separate RNAs.83. The method of any of embodiments 58-82, which does not comprisecontacting the mammalian cell with a template DNA.84. A method of modifying the genome of a human cell, comprisingcontacting the cell with:

(a) a polypeptide or a nucleic acid encoding a polypeptide, wherein thepolypeptide comprises (i) a reverse transcriptase domain, (ii) anendonuclease domain, and optionally (iii) a DNA-binding domain; and

(b) a template RNA (or DNA encoding the template RNA) comprising (i) asequence that binds the polypeptide and (ii) a heterologous objectsequence,

wherein the method results in insertion of the heterologous objectsequence into the human cell's genome,

wherein the human cell does not show upregulation of any DNA repairgenes and/or tumor suppressor genes, or wherein no DNA repair geneand/or tumor suppressor gene is upregulated by more than 10%, 5%, 2%, or1%, e.g., wherein upregulation is measured by RNA-seq, e.g., asdescribed in Example 14.

85. A method of adding an exogenous coding region to the genome of acell (e.g., a mammalian cell), comprising contacting the cell with anRNA comprising the non-coding strand of the exogenous coding region,wherein optionally the RNA does not comprise a coding strand of theexogenous coding region, wherein optionally the delivery comprisesnon-viral delivery.86. A method of expressing a polypeptide in a cell (e.g., a mammaliancell), comprising comprising contacting the cell with an RNA, whereinthe RNA comprises a non-coding strand that is the reverse complement ofa sequence that would encoding the polypeptide, wherein optionally theRNA does not comprise a coding strand encoding the polypeptide, whereinoptionally the delivery comprises non-viral delivery.87. The method of any of embodiments 58-86, wherein the sequence that isinserted into the mammalian genome is a sequence that is exogenous tothe mammalian genome.88. The method of any of embodiments 58-87, which operates independentlyof a DNA template.89. The method of any of embodiments 58-88, wherein the cell is part ofa tissue.90. The method of any of embodiments 58-89, wherein the mammalian cellis euploid, is not immortalized, is part of an organism, is a primarycell, is non-dividing, is a hepatocyte, or is from a subject having agenetic disease.91. The method of any of embodiments 58-90, wherein the contactingcomprises contacting the cell with a plasmid, virus, viral-likeparticle, virosome, liposome, vesicle, exosome, or lipid nanoparticle.92. The method of any of embodiments 58-91, wherein the contactingcomprises using non-viral delivery.93. The method of any of embodiments 58-92, which comprises comprisingcontacting the cell with the template RNA (or DNA encoding the templateRNA), wherein the template RNA comprises the non-coding strand of anexogenous coding region, wherein optionally the template RNA does notcomprise a coding strand of the exogenous coding region, whereinoptionally the delivery comprises non-viral delivery, thereby adding theexogenous coding region to the genome of the cell.94. The method of any of embodiments 58-93, which comprises contactingthe cell with the template RNA (or DNA encoding the template RNA),wherein the template RNA comprises a non-coding strand that is thereverse complement of a sequence that would encoding the polypeptide,wherein optionally the template RNA does not comprise a coding strandencoding the polypeptide, wherein optionally the delivery comprisesnon-viral delivery, thereby expressing the polypeptide in the cell.95. The method of any of embodiments 63-94, wherein the contactingcomprises administering (a) and (b) to a subject, e.g., intravenously.96. The method of any of embodiments 63-95, wherein the contactingcomprises administering a dose of (a) and (b) to a subject at leasttwice.97. The method of any of embodiments 63-96, wherein the polypeptidereverse transcribes the template RNA sequence into the target DNAstrand, thereby modifying the target DNA strand.98. The method of any embodiments 63-97, wherein (a) and (b) areadministered separately.99. The method of any of embodiments 63-97, wherein (a) and (b) areadministered together.100. The method of any of embodiments 63-99, wherein the nucleic acid of(a) is not integrated into the genome of the host cell.101. Any preceding numbered method, wherein the sequence that binds thepolypeptide has one or more of the following characteristics:

(a) is at the 3′ end of the template RNA;

(b) is at the 5′ end of the template RNA;

(b) is a non-coding sequence;

(c) is a structured RNA; or

(d) forms at least 1 hairpin loop structures.

102. Any preceding numbered method, wherein the template RNA furthercomprises a sequence comprising at least 20 nucleotides of at least 80%identity (e.g., at least 85%, 90%, 95%, 97%, 98%, 99%, 100% identity) toa target DNA strand.103. Any preceding numbered method, wherein the template RNA furthercomprises a sequence comprising at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11,12, 13, 14, 15, 20, 25, 30, 35, 40, 45, 50, 60, 70, 80, 90, 100, 110,120, 130, 140, 150 nucleotides of at least 80% identity (e.g., at least85%, 90%, 95%, 97%, 98%, 99%, 100% identity) to a target DNA strand.104. Any preceding numbered method, wherein the sequence comprising atleast 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 20, 25, 30, 35,40, 45, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150 nucleotides, orabout: 2-10, 10-20, 20-30, 30-40, 40-50, 50-60, 60-70, 70-80, 80-90,90-100, 10-100, or 2-100 nucleotides, of at least 80% identity to atarget DNA strand is at the 3′ end of the template RNA.105. Any preceding numbered method, wherein the template RNA furthercomprises a sequence comprising at least 100 nucleotides of at least 80%identity (e.g., at least 85%, 90%, 95%, 97%, 98%, 99%, 100% identity) toa target DNA strand, e.g., at the 3′ end of the template RNA.106. The method of embodiment 104 or 105, wherein the site in the targetDNA strand to which the sequence comprises at least 80% identity isproximal to (e.g., within about: 0-10, 10-20, 20-30, 30-50, or 50-100nucleotides of) a target site on the target DNA strand that isrecognized (e.g., bound and/or cleaved) by the polypeptide comprisingthe endonuclease.107. Any preceding numbered method, wherein the sequence comprising atleast 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 20, 25, 30, 35,40, 45, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150 nucleotides, orabout: 2-10, 10-20, 20-30, 30-40, 40-50, 50-60, 60-70, 70-80, 80-90,90-100, 10-100, or 2-100 nucleotides, of at least 80% identity to atarget DNA strand is at the 3′ end of the template RNA;

optionally wherein the site in the target DNA strand to which thesequence comprises at least 80% identity is proximal to (e.g., withinabout: 0-10, 10-20, or 20-30 nucleotides of) a target site on the targetDNA strand that is recognized (e.g., bound and/or cleaved) by thepolypeptide comprising the endonuclease.

108. The method of embodiment 107, wherein the target site is the sitein the human genome that has the closest identity to a native targetsite of the polypeptide comprising the endonuclease, e.g., wherein thetarget site in the human genome has at least about: 16, 17, 18, 19, or20 nucleotides identical to the native target site.109. Any preceding numbered method, wherein the template RNA has atleast 3, 4, 5, 6, 7, 8, 9, or 10 bases of 100% identity to the targetDNA strand.110. Any preceding numbered method, wherein the at least 3, 4, 5, 6, 7,8, 9, or 10 bases of 100% identity to the target DNA strand are at the3′ end of the template RNA.111. Any preceding numbered method, wherein the at least 3, 4, 5, 6, 7,8, 9, or 10 bases of 100% identity to the target DNA strand are at the5′ end of the template RNA.112. Any preceding numbered method, wherein the template RNA comprisesat least 3, 4, 5, 6, 7, 8, 9, or 10 bases of 100% identity to the targetDNA strand at the 5′ end of the template RNA and at least 3, 4, 5, 6, 7,8, 9, or 10 bases of 100% identity to the target DNA strand at the 3′end of the template RNA.113. Any preceding numbered method, wherein the heterologous objectsequence is between 50-50,000 base pairs (e.g., between 50-40,000 bp,between 500-30,000 bp between 500-20,000 bp, between 100-15,000 bp,between 500-10,000 bp, between 50-10,000 bp, between 50-5,000 bp).114. Any preceding numbered method, wherein the heterologous objectsequence is at least 10, 25, 50, 100, 150, 200, 250, 300, 400, 500, 600,or 700 bp.115. Any preceding numbered method, wherein the heterologous objectsequence is at least 715, 750, 800, 950, 1,000, 2,000, 3,000, or 4,000bp.116. Any preceding numbered method, wherein the heterologous objectsequence is less than 5,000, 10,000, 15,000, 20,000, 30,000, or 40,000bp.117. Any preceding numbered method, wherein the heterologous objectsequence is less than 700, 600, 500, 400, 300, 200, 150, or 100 bp.

118. Any preceding numbered method, wherein the heterologous objectsequence comprises:

(a) an open reading frame, e.g., a sequence encoding a polypeptide,e.g., an enzyme (e.g., a lysosomal enzyme), a membrane protein, a bloodfactor, an exon, an intracellular protein (e.g., a cytoplasmic protein,a nuclear protein, an organellar protein such as a mitochondrial proteinor lysosomal protein), an extracellular protein, a structural protein, asignaling protein, a regulatory protein, a transport protein, a sensoryprotein, a motor protein, a defense protein, or a storage protein;

(b) a non-coding and/or regulatory sequence, e.g., a sequence that bindsa transcriptional modulator, e.g., a promoter, an enhancer, aninsulator;

(c) a splice acceptor site;

(d) a polyA site;

(e) an epigenetic modification site; or

(f) a gene expression unit.

119. Any preceding numbered method, wherein the target DNA is a genomicsafe harbor (GSH) site.120. Any preceding numbered method, wherein the target DNA is a genomicNatural Harbor™ site.121. Any preceding numbered method, which results in insertion of theheterologous object sequence into the a target site in the genome at anaverage copy number of at least 0.01, 0.025, 0.05, 0.075, 0.1, 0.15,0.2, 0.25, 0.3, 0.4, 0.5, 0.75, 1, 1.25, 1.5, 1.75, 2, 2.5, 3, 4, or 5copies per genome.122. Any preceding numbered method, which results in about 25-100%,50-100%, 60-100%, 70-100%, 75-95%, 80%-90%, of integrants into a targetsite in the genome being non-truncated, as measured by an assaydescribed herein, e.g., an assay of Example 6.123. Any preceding numbered method, which results in insertion of theheterologous object sequence only at one target site in the genome ofthe cell.124. Any preceding numbered method, which results in insertion of theheterologous object sequence into a target site in a cell, wherein theinserted heterologous sequence comprises less than 10%, 5%, 2%, 1%,0.5%, 0.2%, or 0.1% mutations (e.g., SNPs or one or more deletions,e.g., truncations or internal deletions) relative to the heterologoussequence prior to insertion, e.g., as measured by an assay of Example12.125. Any preceding numbered method, which results in insertion of theheterologous object sequence into a target site in a plurality of cells,wherein less than 10%, 5%, 2%, or 1% of copies of the insertedheterologous sequence comprise a mutation (e.g., a SNP or a deletion,e.g., a truncation or an internal deletion), e.g., as measured by anassay of Example 12.126. Any preceding numbered method, which results in insertion of theheterologous object sequence into a target cell genome, and wherein thetarget cell does not show upregulation of p53, or shows upregulation ofp53 by less than 10%, 5%, 2%, or 1%, e.g., wherein upregulation of p53is measured by p53 protein level, e.g., according to the methoddescribed in Example 30, or by the level of p53 phosphorylated at Ser15and Ser20.127. Any preceding numbered method, which results in insertion of theheterologous object sequence into a target cell genome, and wherein thetarget cell does not show upregulation of any DNA repair genes and/ortumor suppressor genes, or wherein no DNA repair gene and/or tumorsuppressor gene is upregulated by more than 10%, 5%, 2%, or 1%, e.g.,wherein upregulation is measured by RNA-seq, e.g., as described inExample 14.128. Any preceding numbered method, which results in insertion of theheterologous object sequence into the target site (e.g., at a copynumber of 1 insertion or more than one insertion) in about 1-80% ofcells in a population of cells contacted with the system, e.g., about:1-10%, 10-20%, 20-30%, 30-40%, 40-50%, 50-60%, 60-70%, or 70-80% ofcells, e.g., as measured using single cell ddPCR, e.g., as described inExample 17.129. Any preceding numbered method, which results in insertion of theheterologous object sequence into the target site at a copy number of 1insertion in about 1-80% of cells in a population of cells contactedwith the system, e.g., about: 1-10%, 10-20%, 20-30%, 30-40%, 40-50%,50-60%, 60-70%, or 70-80% of cells, e.g., as measured using colonyisolation and ddPCR, e.g., as described in Example 18.130. Any preceding numbered method, which results in insertion of theheterologous object sequence into the target site (on-target insertions)at a higher rate that insertion into a non-target site (off-targetinsertions) in a population of cells, wherein the ratio of on-targetinsertions to off-target insertions is greater than 10:1, 20:1, 30:1,40:1, 50:1, 60:1, 70:1, 80:1. 90:1, 100:1, 200:1, 500:1, or 1,000:1,e.g., using an assay of Example 11.131. Any above-numbered method, results in insertion of a heterologousobject sequence in the presence of an inhibitor of a DNA repair pathway(e.g., SCR7, a PARP inhibitor), or in a cell line deficient for a DNArepair pathway (e.g., a cell line deficient for the nucleotide excisionrepair pathway or the homology-directed repair pathway).132. Any preceding numbered system, formulated as a pharmaceuticalcomposition.133. Any preceding numbered system, disposed in a pharmaceuticallyacceptable carrier (e.g., a vesicle, a liposome, a natural or syntheticlipid bilayer, a lipid nanoparticle, an exosome).134. A method of making a system for modifying the genome of a mammaliancell, comprising:

a) providing a template RNA as described in any of the precedingembodiments, e.g., wherein the template RNA comprises (i) a sequencethat binds a polypeptide comprising a reverse transcriptase domain andan endonuclease domain, and (ii) a heterologous object sequence; and

b) treating the template RNA to reduce secondary structure, e.g.,heating the template RNA, e.g., to at least 70, 75, 80, 85, 90, or 95 C,and

c) subsequently cooling the template RNA, e.g., to a temperature thatallows for secondary structure, e.g, to less than or equal to 30, 25, or20 C.

135. The method of embodiment 134, which further comprises contactingthe template RNA with a polypeptide that comprises (i) a reversetranscriptase domain and (ii) an endonuclease domain, or with a nucleicacid (e.g., RNA) encoding the polypeptide.136. The method of embodiment 134 or 135, which further comprisescontacting the template RNA with a cell.137. The system or method of any of the preceding embodiments, whereinthe heterologous object sequence encodes a therapeutic polypeptide.138. The system or method of any of the preceding embodiments, whereinthe heterologous object sequence encodes a mammalian (e.g., human)polypeptide, or a fragment or variant thereof.139. The system or method of any of the preceding embodiments, whereinthe heterologous object sequence encodes an enzyme (e.g., a lysosomalenzyme), a blood factor (e.g., Factor I, II, V, VII, X, XI, XII orXIII), a membrane protein, an exon, an intracellular protein (e.g., acytoplasmic protein, a nuclear protein, an organellar protein such as amitochondrial protein or lysosomal protein), an extracellular protein, astructural protein, a signaling protein, a regulatory protein, atransport protein, a sensory protein, a motor protein, a defenseprotein, or a storage protein.140. The system or method of any of the preceding embodiments, whereinthe heterologous object sequence comprises a tissue specific promoter orenhancer.141. The system or method of any of the preceding embodiments, whereinthe heterologous object sequence encodes a polypeptide of greater than250, 300, 400, 500, or 1,000 amino acids, and optionally up to 1300amino acids.142. The system or method of any of the preceding embodiments, whereinthe heterologous object sequence encodes a fragment of a mammalian genebut does not encode the full mammalian gene, e.g., encodes one or moreexons but does not encode a full-length protein.143. The system or method of any of the preceding embodiments, whereinthe heterologous object sequence encodes one or more introns.144. The system or method of any of the preceding embodiments, whereinthe heterologous object sequence is other than a GFP, e.g., is otherthan a fluorescent protein or is other than a reporter protein.145. The system or method of any of the preceding embodiments, whereinthe polypeptide comprises (i) a reverse transcriptase domain and (ii) anendonuclease domain, wherein one or both of (i) or (ii) are derived froman avian retrotransposase, e.g., have a sequence of Table 2 or 3 or atleast 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% identitythereto.146. The system or method of any of the preceding embodiments, whereinthe polypeptide has an activity at 37° C. that is no less than 70%, 75%,80%, 85%, 90%, or 95% of its activity at 25° C. under otherwise similarconditions.147. The system or method of any of the preceding embodiments, whereinthe nucleic acid encoding the polypeptide and the template RNA or anucleic acid encoding the template RNA are separate nucleic acids.148. The system or method of any of the preceding embodiments, whereinthe template RNA does not encode an active reverse transcriptase, e.g.,comprises an inactivated mutant reverse transcriptase, e.g., asdescribed in Example 1 or 2, or does not comprise a reversetranscriptase sequence.149. The system or method of any of the preceding embodiments, whereinthe template RNA comprises one or more chemical modifications.150. The system or method of any of the preceding embodiments, whereinthe heterologous object sequence is disposed between the promoter andthe sequence that binds the polypeptide.151. The system or method of any of the preceding embodiments, whereinthe promoter is disposed between the heterologous object sequence andthe sequence that binds the polypeptide.152. The system or method of any of the preceding embodiments, whereinthe heterologous object sequence comprises an open reading frame (or thereverse complement thereof) in a 5′ to 3′ orientation on the templateRNA.153. The system or method of any of the preceding embodiments, whereinthe heterologous object sequence comprises an open reading frame (or thereverse complement thereof) in a 3′ to 5′ orientation on the templateRNA.154. The system or method of any of the preceding embodiments, whereinthe polypeptide comprises (a) a reverse transcriptase domain and (b) anendonuclease domain, wherein at least one of (a) or (b) is heterologous.155. The system or method of any of the preceding embodiments, whereinthe polypeptide comprises (a) a target DNA binding domain, (b) a reversetranscriptase domain and (c) an endonuclease domain, wherein at leastone of (a), (b) or (c) is heterologous.156. A substantially pure polypeptide comprising (a) a reversetranscriptase domain and (b) a heterologous endonuclease domain.157. A substantially pure polypeptide comprising (a) a target DNAbinding domain, (b) a reverse transcriptase domain and (c) anendonuclease domain, wherein at least one of (a), (b) or (c) isheterologous.158. A substantially pure polypeptide comprising (a) a reversetranscriptase domain, (b) an endonuclease domain, and (c) a heterologoustarget DNA binding domain.159. A polypeptide or a nucleic acid encoding the polypeptide, whereinthe polypeptide comprises (a) a reverse transcriptase domain and (b) anendonuclease domain, wherein at least one of (a) or (b) is heterologousto the other.160. A polypeptide or a nucleic acid encoding the polypeptide, whereinthe polypeptide comprises (a) a target DNA binding domain, (b) a reversetranscriptase domain and (c) an endonuclease domain, wherein at leastone of (a), (b) or (c) is heterologous to the other.161. Any polypeptide of numbered embodiments 156-160, wherein thereverse transcriptase domain has at least 80% identity (e.g., at least85%, 90%, 95%, 97%, 98%, 99%, 100% identity) to a reverse transcriptasedomain of an APE-type or RLE-type non-LTR retrotransposon listed in anyof Tables 1-3.162. Any polypeptide of numbered embodiments 156-161, wherein theendonuclease domain has at least 80% identity e.g., at least 85%, 90%,95%, 97%, 98%, 99%, 100% identity, to a endonuclease domain of anAPE-type or RLE-type non-LTR retrotransposon listed in any of Tables1-3.163. Any polypeptide of numbered embodiments 156-162 or any precedingnumbered method, wherein the DNA binding domain has at least 80%identity e.g., at least 85%, 90%, 95%, 97%, 98%, 99%, 100% identity, toa DNA binding domain of a sequence listed in Table 1, 2, or 3.164. A nucleic acid encoding the polypeptide of any preceding numberedembodiment.165. A vector comprising the nucleic acid of numbered embodiment 164.166. A host cell comprising the nucleic acid of numbered embodiment 164.167. A host cell comprising the polypeptide of any preceding numberedembodiment.168. A host cell comprising the vector of numbered embodiment 165.169. A host cell (e.g., a human cell) comprising: (i) a heterologousobject sequence (e.g., a sequence encoding a therapeutic polypeptide) ata target site in a chromosome, and (ii) one or both of an untranslatedregion (e.g., a retrotransposon untranslated sequence, e.g., a sequenceof column 6 of Table 3) on one side (e.g., upstream) of the heterologousobject sequence, and an untranslated region (e.g., a retrotransposonuntranslated sequence, e.g., a sequence of column 7 of Table 3) on theother side (e.g., downstream) of the heterologous object sequence.170. A host cell (e.g., a human cell) comprising: (i) a heterologousobject sequence (e.g., a sequence encoding a therapeutic polypeptide) ata target site in a chromosome, wherein the target locus is a NaturalHarbor™ site, e.g., a site of Table 4 herein.171. The host cell of embodiment 170, which further comprises (ii) oneor both of an untranslated region 5′ of the heterologous objectsequence, and an untranslated region 3′ of the heterologous objectsequence.172. The host cell of embodiment 170, which further comprises (ii) oneor both of an untranslated region (e.g., a retrotransposon untranslatedsequence, e.g., a sequence of column 6 of Table 3) on one side (e.g.,upstream) of the heterologous object sequence, and an untranslatedregion (e.g., a retrotransposon untranslated sequence, e.g., a sequenceof column 7 of Table 3) on the other side (e.g., downstream) of theheterologous object sequence.173. The host cell of any of embodiments 169-173, which comprisesheterologous object sequence at only the target site.174. A pharmaceutical composition, comprising any preceding numberedsystem, nucleic acid, polypeptide, or vector; and a pharmaceuticallyacceptable excipient or carrier.175. The pharmaceutical composition of embodiment 174, wherein thepharmaceutically acceptable excipient or carrier is selected from avector (e.g., a viral or plasmid vector), a vesicle (e.g., a liposome,an exosome, a natural or synthetic lipid bilayer), a lipid nanoparticle.176. A polypeptide of any of the preceding embodiments, wherein thepolypeptide further comprises a nuclear localization sequence.177. A method of modifying a target DNA strand in a cell, tissue orsubject, comprising administering any preceding numbered system to thecell, tissue or subject, thereby modifying the target DNA strand.178. Any preceding numbered embodiment, wherein the polypeptidecomprises an amino acid sequence having at least 70%, 75%, 80%, 85%,90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an amino acidsequence listed in Table 5 (e.g., any one of SEQ ID NOs: 1017-1022), ora functional fragment thereof.179. Any preceding numbered embodiment, wherein the reversetranscriptase domain comprises an amino acid sequence having at least70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequenceidentity to the reverse transcriptase domain of an amino acid sequencelisted in Table 5 (e.g., any one of SEQ ID NOs: 1017-1022), or afunctional fragment thereof.180. Any preceding numbered embodiment, wherein the retrotransposasecomprises an amino acid sequence having at least 70%, 75%, 80%, 85%,90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to an amino acidsequence listed in Table 5 (e.g., any one of SEQ ID NOs: 1017-1022), ora functional fragment thereof.181. Any preceding numbered embodiment, wherein the polypeptidecomprises an amino acid sequence having at least 70%, 75%, 80%, 85%,90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the aminoacid sequence SGSETPGTSESATPES (SEQ ID NO: 1023) or GGGS (SEQ ID NO:1024).182. Any preceding numbered embodiment, wherein the reversetranscriptase domain comprises an amino acid sequence having at least70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequenceidentity to the amino acid sequence SGSETPGTSESATPES (SEQ ID NO: 1023)or GGGS (SEQ ID NO: 1024).183. Any preceding numbered embodiment, wherein the retrotransposasecomprises an amino acid sequence having at least 70%, 75%, 80%, 85%,90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the aminoacid sequence SGSETPGTSESATPES (SEQ ID NO: 1023) or GGGS (SEQ ID NO:1024).184. Any preceding numbered embodiment, wherein the polypeptide, reversetranscriptase domain, or retrotransposase comprises a linker comprisingan amino acid sequence having at least 70%, 75%, 80%, 85%, 90%, 95%,96%, 97%, 98%, 99%, or 100% sequence identity to the amino acid sequenceSGSETPGTSESATPES (SEQ ID NO: 1023) or GGGS (SEQ ID NO: 1024).185. Any preceding numbered embodiment, wherein the polypeptidecomprises a DNA binding domain covalently attached to the remainder ofthe polypeptide by a linker, e.g., a linker comprising at least 1, 2, 3,4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 25, 30,35, 40, 45, 50, 60, 70, 80, 90, 100, 125, 150, 200, 300, 400, or 500amino acids.186. Numbered embodiment 185, wherein the linker is attached to theremainder of the polypeptide at a position in the DNA binding domain,RNA binding domain, reverse transcriptase domain, or endonuclease domain(e.g., as shown in any of FIGS. 17A-17F).187. Numbered embodiment 185 or 186, wherein the linker is attached tothe remainder of the polypeptide at a position in the N-terminal side ofan alpha helical region of the polypeptide, e.g., at a positioncorresponding to version v1 as described in Example 26.188. Numbered embodiment 185 or 186, wherein the linker is attached tothe remainder of the polypeptide at a position in the C-terminal side ofan alpha helical region of the polypeptide, e.g., preceding an RNAbinding motif (e.g., a −1 RNA binding motif), e.g., at a positioncorresponding to version v2 as described in Example 26.189. Numbered embodiment 185 or 186, wherein the linker is attached tothe remainder of the polypeptide at a position in the C-terminal side ofa random coil region of the polypeptide, e.g., N-terminal relative to aDNA binding motif (e.g., a c-myb DNA binding motif), e.g., at a positioncorresponding to version v3 as described in Example 26.190. Any one of numbered embodiments 185-189, wherein the linkercomprises an amino acid sequence having at least 70%, 75%, 80%, 85%,90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the aminoacid sequence SGSETPGTSESATPES (SEQ ID NO: 1023) or GGGS (SEQ ID NO:1024).191. Any preceding numbered embodiment, wherein a polynucleotidesequence comprising at least about 500, 1000, 2000, 3000, 3500, 3600,3700, 3800, 3900, or 4000 contiguous nucleotides from the 5′ end of thetemplate RNA sequence are integrated into a target cell genome.192. Any preceding numbered embodiment, wherein a polynucleotidesequence comprising at least about 500, 1000, 2000, 2500, 2600, 2700,2800, 2900, or 3000 contiguous nucleotides from the 3′ end of thetemplate RNA sequence are integrated into a target cell genome.193. Any preceding numbered embodiment, wherein the nucleic acidsequence of the template RNA, or a portion thereof (e.g., a portioncomprising at least about 100, 200, 300, 400, 500, 1000, 2000, 2500,3000, 3500, or 4000 nucleotides) integrates into the genomes of apopulation of target cells at a copy number of at least about 0.21, 0.3,0.4, 0.5, 0.6, 0.7, 0.8, 0.9, or 1.0 integrants/genome.194. Any preceding numbered embodiment, wherein the nucleic acidsequence of the template RNA, or a portion thereof (e.g., a portioncomprising at least about 100, 200, 300, 400, 500, 1000, 2000, 2500,3000, 3500, or 4000 nucleotides) integrates into the genomes of apopulation of target cells at a copy number of at least about 0.085,0.09, 0.1, 0.15, or 0.2 integrants/genome.195. Any preceding numbered embodiment, wherein the nucleic acidsequence of the template RNA, or a portion thereof (e.g., a portioncomprising at least about 100, 200, 300, 400, 500, 1000, 2000, 2500,3000, 3500, or 4000 nucleotides) integrates into the genomes of apopulation of target cells at a copy number of at least about 0.036,0.04, 0.05, 0.06, 0.07, or 0.08 integrants/genome.196. Any preceding numbered embodiment, wherein the polypeptidecomprises a functional endonuclease domain (e.g., wherein theendonuclease domain does not comprise a mutation that abolishesendonuclease activity, e.g., as described herein).197. Any preceding numbered embodiment, wherein the polypeptidecomprises an amino acid sequence having at least 70%, 75%, 80%, 85%,90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the R2polypeptide from a medium ground finch, e.g., Geospiza fortis (e.g., asdescribed herein, e.g., R2-1_GFo), or a functional fragment thereof.198. Any preceding numbered embodiment, wherein the reversetranscriptase domain comprises an amino acid sequence having at least70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequenceidentity to the R2 polypeptide from a medium ground finch, e.g.,Geospiza fortis (e.g., as described herein, e.g., R2-1_GFo), or afunctional fragment thereof.199. Any preceding numbered embodiment, wherein the retrotransposasecomprises an amino acid sequence having at least 70%, 75%, 80%, 85%,90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the R2polypeptide from a medium ground finch, e.g., Geospiza fortis (e.g., asdescribed herein, e.g., R2-1_GFo), or a functional fragment thereof.200. Any one of numbered embodiments 197-199, wherein the nucleic acidsequence of the template RNA, or a portion thereof (e.g., a portioncomprising at least about 100, 200, 300, 400, 500, 1000, 2000, 2500,3000, 3500, or 4000 nucleotides) integrates into the genomes of apopulation of target cells at a copy number of at least about 0.21integrants/genome.201. Any preceding numbered embodiment, wherein the polypeptidecomprises an amino acid sequence having at least 70%, 75%, 80%, 85%,90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the R4polypeptide from a large roundworm, e.g., Ascaris lumbricoides (e.g., asdescribed herein, e.g., R4_AL), or a functional fragment thereof.202. Any preceding numbered embodiment, wherein the reversetranscriptase domain comprises an amino acid sequence having at least70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% sequenceidentity to the R4 polypeptide from a large roundworm, e.g., Ascarislumbricoides (e.g., as described herein, e.g., R4_AL), or a functionalfragment thereof.203. Any preceding numbered embodiment, wherein the retrotransposasecomprises an amino acid sequence having at least 70%, 75%, 80%, 85%,90%, 95%, 96%, 97%, 98%, 99%, or 100% sequence identity to the R4polypeptide from a large roundworm, e.g., Ascaris lumbricoides (e.g., asdescribed herein, e.g., R4_AL), or a functional fragment thereof.204. Any one of numbered embodiments 201-203, wherein the nucleic acidsequence of the template RNA, or a portion thereof (e.g., a portioncomprising at least about 100, 200, 300, 400, 500, 1000, 2000, 2500,3000, 3500, or 4000 nucleotides) integrates into the genomes of apopulation of target cells at a copy number of at least about 0.085integrants/genome.205. Any preceding numbered embodiment, wherein introduction of thesystem into a target cell does not result in alteration (e.g.,upregulation) of p53 and/or p21 protein levels, H2AX phosphorylation(e.g., gamma H2AX), ATM phosphorylation, ATR phosphorylation, Chk1phosphorylation, Chk2 phosphorylation, and/or p53 phosphorylation.206. Any preceding numbered embodiment, wherein introduction of thesystem into a target cell results in upregulation of p53 protein levelin the target cell to a level that is less than about 0.001%, 0.005%,0.01%, 0.05%, 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%,20%, 30%, 40%, 45%, 50%, 55%, 60%, 70%, 80%, or 90% of the p53 proteinlevel induced by introducing a site-specific nuclease, e.g., Cas9, thattargets the same genomic site as said system.207. Numbered embodiment 205 or 206, wherein the p53 protein level isdetermined according to the method described in Example 30.208. Any preceding numbered embodiment, wherein introduction of thesystem into a target cell results in upregulation of p53 phosphorylationlevel in the target cell to a level that is less than about 0.001%,0.005%, 0.01%, 0.05%, 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%,10%, 15%, 20%, 30%, 40%, 45%, 50%, 55%, 60%, 70%, 80%, or 90% of the p53phosphorylation level induced by introducing a site-specific nuclease,e.g., Cas9, that targets the same genomic site as said system.209. Any preceding numbered embodiment, wherein introduction of thesystem into a target cell results in upregulation of p21 protein levelin the target cell to a level that is less than about 0.001%, 0.005%,0.01%, 0.05%, 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 15%,20%, 30%, 40%, 45%, 50%, 55%, 60%, 70%, 80%, or 90% of the p53 proteinlevel induced by introducing a site-specific nuclease, e.g., Cas9, thattargets the same genomic site as said system.210. Numbered embodiment 205 or 209, wherein the p21 protein level isdetermined according to the method described in Example 30.211. Any preceding numbered embodiment, wherein introduction of thesystem into a target cell results in upregulation of H2AXphosphorylation level in the target cell to a level that is less thanabout 0.001%, 0.005%, 0.01%, 0.05%, 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%,7%, 8%, 9%, 10%, 15%, 20%, 30%, 40%, 45%, 50%, 55%, 60%, 70%, 80%, or90% of the H2AX phosphorylation level induced by introducing asite-specific nuclease, e.g., Cas9, that targets the same genomic siteas said system.212. Any preceding numbered embodiment, wherein introduction of thesystem into a target cell results in upregulation of ATM phosphorylationlevel in the target cell to a level that is less than about 0.001%,0.005%, 0.01%, 0.05%, 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%,10%, 15%, 20%, 30%, 40%, 45%, 50%, 55%, 60%, 70%, 80%, or 90% of the ATMphosphorylation level induced by introducing a site-specific nuclease,e.g., Cas9, that targets the same genomic site as said system.213. Any preceding numbered embodiment, wherein introduction of thesystem into a target cell results in upregulation of ATR phosphorylationlevel in the target cell to a level that is less than about 0.001%,0.005%, 0.01%, 0.05%, 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%,10%, 15%, 20%, 30%, 40%, 45%, 50%, 55%, 60%, 70%, 80%, or 90% of the ATRphosphorylation level induced by introducing a site-specific nuclease,e.g., Cas9, that targets the same genomic site as said system.214. Any preceding numbered embodiment, wherein introduction of thesystem into a target cell results in upregulation of Chk1phosphorylation level in the target cell to a level that is less thanabout 0.001%, 0.005%, 0.01%, 0.05%, 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%,7%, 8%, 9%, 10%, 15%, 20%, 30%, 40%, 45%, 50%, 55%, 60%, 70%, 80%, or90% of the Chk1 phosphorylation level induced by introducing asite-specific nuclease, e.g., Cas9, that targets the same genomic siteas said system.215. Any preceding numbered embodiment, wherein introduction of thesystem into a target cell results in upregulation of Chk2phosphorylation level in the target cell to a level that is less thanabout 0.001%, 0.005%, 0.01%, 0.05%, 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%,7%, 8%, 9%, 10%, 15%, 20%, 30%, 40%, 45%, 50%, 55%, 60%, 70%, 80%, or90% of the Chk2 phosphorylation level induced by introducing asite-specific nuclease, e.g., Cas9, that targets the same genomic siteas said system.

Definitions

Domain: The term “domain” as used herein refers to a structure of abiomolecule that contributes to a specified function of the biomolecule.A domain may comprise a contiguous region (e.g., a contiguous sequence)or distinct, non-contiguous regions (e.g., non-contiguous sequences) ofa biomolecule. Examples of protein domains include, but are not limitedto, an endonuclease domain, a DNA binding domain, a reversetranscription domain; an example of a domain of a nucleic acid is aregulatory domain, such as a transcription factor binding domain.

Exogenous: As used herein, the term exogenous, when used with referenceto a biomolecule (such as a nucleic acid sequence or polypeptide) meansthat the biomolecule was introduced into a host genome, cell or organismby the hand of man. For example, a nucleic acid that is as added into anexisting genome, cell, tissue or subject using recombinant DNAtechniques or other methods is exogenous to the existing nucleic acidsequence, cell, tissue or subject.

Genomic safe harbor site (GSH site): A genomic safe harbor site is asite in a host genome that is able to accommodate the integration of newgenetic material, e.g., such that the inserted genetic element does notcause significant alterations of the host genome posing a risk to thehost cell or organism. A GSH site generally meets 1, 2, 3, 4, 5, 6, 7, 8or 9 of the following criteria: (i) is located >300 kb from acancer-related gene; (ii) is >300 kb from a miRNA/other functional smallRNA; (iii) is >50 kb from a 5′ gene end; (iv) is >50 kb from areplication origin; (v) is >50 kb away from any ultraconserveredelement; (vi) has low transcriptional activity (i.e. no mRNA+/−25 kb);(vii) is not in copy number variable region; (viii) is in openchromatin; and/or (ix) is unique, with 1 copy in the human genome.Examples of GSH sites in the human genome that meet some or all of thesecriteria include (i) the adeno-associated virus site 1 (AAVS1), anaturally occurring site of integration of AAV virus on chromosome 19;(ii) the chemokine (C-C motif) receptor 5 (CCR5) gene, a chemokinereceptor gene known as an HIV-1 coreceptor; (iii) the human ortholog ofthe mouse Rosa26 locus; (iv) the rDNA locus. Additional GSH sites areknown and described, e.g., in Pellenz et al. epub Aug. 20, 2018(https://doi.org/10.1101/396390).

Heterologous: The term heterologous, when used to describe a firstelement in reference to a second element means that the first elementand second element do not exist in nature disposed as described. Forexample, a heterologous polypeptide, nucleic acid molecule, construct orsequence refers to (a) a polypeptide, nucleic acid molecule or portionof a polypeptide or nucleic acid molecule sequence that is not native toa cell in which it is expressed, (b) a polypeptide or nucleic acidmolecule or portion of a polypeptide or nucleic acid molecule that hasbeen altered or mutated relative to its native state, or (c) apolypeptide or nucleic acid molecule with an altered expression ascompared to the native expression levels under similar conditions. Forexample, a heterologous regulatory sequence (e.g., promoter, enhancer)may be used to regulate expression of a gene or a nucleic acid moleculein a way that is different than the gene or a nucleic acid molecule isnormally expressed in nature. In another example, a heterologous domainof a polypeptide or nucleic acid sequence (e.g., a DNA binding domain ofa polypeptide or nucleic acid encoding a DNA binding domain of apolypeptide) may be disposed relative to other domains or may be adifferent sequence or from a different source, relative to other domainsor portions of a polypeptide or its encoding nucleic acid. In certainembodiments, a heterologous nucleic acid molecule may exist in a nativehost cell genome, but may have an altered expression level or have adifferent sequence or both. In other embodiments, heterologous nucleicacid molecules may not be endogenous to a host cell or host genome butinstead may have been introduced into a host cell by transformation(e.g., transfection, electroporation), wherein the added molecule mayintegrate into the host genome or can exist as extra-chromosomal geneticmaterial either transiently (e.g., mRNA) or semi-stably for more thanone generation (e.g., episomal viral vector, plasmid or otherself-replicating vector).

Mutation or Mutated: The term “mutated” when applied to nucleic acidsequences means that nucleotides in a nucleic acid sequence may beinserted, deleted or changed compared to a reference (e.g., native)nucleic acid sequence. A single alteration may be made at a locus (apoint mutation) or multiple nucleotides may be inserted, deleted orchanged at a single locus. In addition, one or more alterations may bemade at any number of loci within a nucleic acid sequence. A nucleicacid sequence may be mutated by any method known in the art.

Nucleic acid molecule: Nucleic acid molecule refers to both RNA and DNAmolecules including, without limitation, cDNA, genomic DNA and mRNA, andalso includes synthetic nucleic acid molecules, such as those that arechemically synthesized or recombinantly produced, such as RNA templates,as described herein. The nucleic acid molecule can be double-stranded orsingle-stranded, circular or linear. If single-stranded, the nucleicacid molecule can be the sense strand or the antisense strand. Unlessotherwise indicated, and as an example for all sequences describedherein under the general format “SEQ. ID NO:,” “nucleic acid comprisingSEQ. ID NO:1” refers to a nucleic acid, at least a portion which haseither (i) the sequence of SEQ. ID NO:1, or (ii) a sequencecomplimentary to SEQ. ID NO:1. The choice between the two is dictated bythe context in which SEQ. ID NO:1 is used. For instance, if the nucleicacid is used as a probe, the choice between the two is dictated by therequirement that the probe be complimentary to the desired target.Nucleic acid sequences of the present disclosure may be modifiedchemically or biochemically or may contain non-natural or derivatizednucleotide bases, as will be readily appreciated by those of skill inthe art. Such modifications include, for example, labels, methylation,substitution of one or more naturally occurring nucleotides with ananalog, inter-nucleotide modifications such as uncharged linkages (forexample, methyl phosphonates, phosphotriesters, phosphoramidates,carbamates, etc.), charged linkages (for example, phosphorothioates,phosphorodithioates, etc.), pendant moieties, (for example,polypeptides), intercalators (for example, acridine, psoralen, etc.),chelators, alkylators, and modified linkages (for example, alphaanomeric nucleic acids, etc.). Also included are synthetic moleculesthat mimic polynucleotides in their ability to bind to a designatedsequence via hydrogen bonding and other chemical interactions. Suchmolecules are known in the art and include, for example, those in whichpeptide linkages substitute for phosphate linkages in the backbone of amolecule. Other modifications can include, for example, analogs in whichthe ribose ring contains a bridging moiety or other structure such asmodifications found in “locked” nucleic acids.

Gene expression unit: a gene expression unit is a nucleic acid sequencecomprising at least one regulatory nucleic acid sequence operably linkedto at least one effector sequence. A first nucleic acid sequence isoperably linked with a second nucleic acid sequence when the firstnucleic acid sequence is placed in a functional relationship with thesecond nucleic acid sequence. For instance, a promoter or enhancer isoperably linked to a coding sequence if the promoter or enhancer affectsthe transcription or expression of the coding sequence. Operably linkedDNA sequences may be contiguous or non-contiguous. Where necessary tojoin two protein-coding regions, operably linked sequences may be in thesame reading frame.

Host: The terms host genome or host cell, as used herein, refer to acell and/or its genome into which protein and/or genetic material hasbeen introduced. It should be understood that such terms are intended torefer not only to the particular subject cell and/or genome, but to theprogeny of such a cell and/or the genome of the progeny of such a cell.Because certain modifications may occur in succeeding generations due toeither mutation or environmental influences, such progeny may not, infact, be identical to the parent cell, but are still included within thescope of the term “host cell” as used herein. A host genome or host cellmay be an isolated cell or cell line grown in culture, or genomicmaterial isolated from such a cell or cell line, or may be a host cellor host genome which composing living tissue or an organism. In someinstances, a host cell may be an animal cell or a plant cell, e.g., asdescribed herein. In certain instances, a host cell may be a bovinecell, horse cell, pig cell, goat cell, sheep cell, chicken cell, orturkey cell. In certain instances, a host cell may be a corn cell, soycell, wheat cell, or rice cell.

Pseudoknot: A “pseudoknot sequence” sequence, as used herein, refers toa nucleic acid (e.g., RNA) having a sequence with suitableself-complementarity to form a pseudoknot structure, e.g., having: afirst segment, a second segment between the first segment and a thirdsegment, wherein the third segment is complementary to the firstsegment, and a fourth segment, wherein the fourth segment iscomplementary to the second segment. The pseudoknot may optionally haveadditional secondary structure, e.g., a stem loop disposed in the secondsegment, a stem-loop disposed between the second segment and thirdsegment, sequence before the first segment, or sequence after the fourthsegment. The pseudoknot may have additional sequence between the firstand second segments, between the second and third segments, or betweenthe third and fourth segments. In some embodiments, the segments arearranged, from 5′ to 3′: first, second, third, and fourth. In someembodiments, the first and third segments comprise five base pairs ofperfect complementarity. In some embodiments, the second and fourthsegments comprise 10 base pairs, optionally with one or more (e.g., two)bulges. In some embodiments, the second segment comprises one or moreunpaired nucleotides, e.g., forming a loop. In some embodiments, thethird segment comprises one or more unpaired nucleotides, e.g., forminga loop.

Stem-loop sequence: As used herein, a “stem-loop sequence” refers to anucleic acid sequence (e.g., RNA sequence) with sufficientself-complementarity to form a stem-loop, e.g., having a stem comprisingat least two (e.g., 3, 4, 5, 6, 7, 8, 9, or 10) base pairs, and a loopwith at least three (e.g., four) base pairs. The stem may comprisemismatches or bulges.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a schematic of the Gene Writing genome editing system.

FIG. 2 is a schematic of the structure of the Gene Writer genome editorpolypeptide.

FIG. 3 is a schematic of a Gene Writer genome editor polypeptidecomprising a heterologous DNA binding domain designed to targetdifferent sites of the genome.

FIG. 4 is a schematic of the structure of Gene Writer genome editortemplate RNA.

FIG. 5 is a schematic showing the Gene Writing genome editing system toadd a gene expression unit into a safe harbor site in the genome.

FIG. 6 is a schematic showing Gene Writing genome editing to add a newexon into an specific intron in the genome and replace downstream exons.

FIG. 7 illustrates a schematic of Miseq library construction. Nested PCRwas performed across the R2Tg-rDNA junction using (1) outer forwardprimer and tailed inner reverse primer followed by (2) tailed innerforward primer and tail reverse primer. The inner reverse primercontains a 1-4 base stagger, an 8-nucleotide randomized UMI, and amultiplexing barcode. The UMI allows for counting of individualamplification events to eliminate PCR bias.

FIGS. 8A-8B: Results of Miseq and Matlab analysis of DNA-mediated R2Tgintegration into Hek293T cells. Each graph shows analysis of (FIG. 8A)the experimental R2Tg (FIG. 8B) and 1 bp deletion negative control. They-axis indicates aligned counts of unique sequences determined viaunique UMIs found via Matlab. The X-axis indicates the sequence positionof sequence coverage. The vertical gray line at the left of the graphindicates the position of the forward primer, while the vertical grayline at the right of the graph indicates the expected Tg-rDNA junctionsite. Bars at the right end of the graph indicate insertion without atruncation, and bars at the left end of the graph indicate truncation.FIG. 8A shows that most sequences show high alignment to the expectedintegration product.

FIG. 9 shows a ddPCR evaluation of copy number variation of theR2Tg-rDNA junction in human cells across transfection conditions.Forward primer and probe were expected to bind to the 3′ UTR of theR2Tg, while reverse primer was targeted to the human rDNA. The resultingddPCR signal was normalized to that of reference assay RPP30 todetermine copy number. Significantly higher average copies per genomewere found with the wildtype (WT, left set of bars) R2Tg as compared togenetic control altering translation with a 1-bp deletion (Frameshiftmutant control, right set of bars).

FIG. 10 illustrates the sequence alignment and coverage of TOPO cloningthe nested PCR product from Example 7. The gray line at the right edgeof the graph indicates the expected transgene-rDNA junction. Mostsequences showed high alignment to the expected integrated product.

FIG. 11 is a schematic of an exemplary template RNA. It comprises apayload domain in the center (e.g., a heterologous object sequence,e.g., comprising a promoter and a protein-coding sequence). The payloaddomain is flanked by 5′ and 3′ protein interaction domains, e.g.,sequences capable of binding the Gene Writer polypeptide, e.g., 5′ and3′ UTR sequences shown in Table 3. Flanking the protein interactiondomains are 5′ and 3′ homology domains, which have homology to thedesired insertion region in the genome.

FIG. 12 is a graph showing retrotransposition efficiency measured byddPCR (digital droplet PCR) using different transfection conditions.Bars A-C represent samples that were transfected using 0.15 μlLipofectamine™ RNAiMAX with 100 ng, 250 ng, or 500 ng respectively. BarsD-F represent samples that were transfected using 0.3 μl Lipofectamine™RNAiMAX with 100 ng, 250 ng, or 500 ng respectively. Bars G-I representsamples that were transfected using 10 TransIT®-mRNA transfection kitwith 100 ng, 250 ng, or 500 ng respectively.

FIG. 13. Schematic of trans-transgene delivery machinery. This schematicillustrates a driver plasmid (left) with a pCEP4 backbone, which encodesthe reverse transcriptase R2Tg, with a promoter and Kozak sequenceupstream, and a polyadenylation signal downstream. The driver plasmidcan drive expression of the GeneWriter protein. The transgene plasmid(right), with a pCDNA backbone, comprises (in order) a CMV promoter, anrDNA homology sequence, a 5′ UTR, an antisense-orientation insert, a 3′UTR, a second rDNA homology sequence, a second polyadenylation signal,and a TK promoter driving a mKate2 marker. The antisense-orientationinsert comprises an EF1α promoter, a coding region for EGFP thatcomprises an intron, and a polyadenylation signal. Use of the CMVpromoter in the transgene plasmid drives expression of a template RNAcomprising the rDNA homology regions, the UTRs, and theantisense-orientation insert.

FIG. 14 shows ddPCR evaluation of copy number variation of thetransgene-rDNA junction in human cells across transfection conditions.Forward primer and probe were designed to bind to the 3′ UTR of theR2Tg, while reverse primer was targeted to the human rDNA. The resultingddPCR signal was normalized to that of reference assay RPP30 todetermine copy number. Significantly higher average copies per genomewere found with the wildtype (WT) R2Tg as compared to backbone constructwith no R2Tg sequence involved. Condition 1 denotes a driver plasmid:transgene plasmid molar ratio of 9:1; condition 2 denotes the ratio is4:1, condition 3 denotes the ratio is 1:1, condition 4 denotes the ratiois 1:4, and condition 5 denotes the ratio is 1:9.

FIGS. 15A and 15B. FIG. 15A: Hybrid capture of R2Tg identified on-targetintegrations in the human genome. The read coverage as aligned to theexpected target integration in the R2 ribosomal site is indicated on they-axis. The 5′ junction between rDNA and R2Tg is indicated by the leftvertical line, while the 3′ junction is indicated by the right verticalline. Next-generation sequencing identifies reads spanning the expectedjunctions. FIG. 15B shows the number of reads from this experimentcategorized as on-target integration or off-target integration at the 5′end and 3′ end of the integrated sequence.

FIG. 16. Sanger sequencing result of the 3′ junction nested PCR.Lowercase nucleotides represent the designed SNP. Shaded uppercasenucleotides represent WT sequence. FIG. 16 discloses SEQ ID NO: 1538.

FIGS. 17A-17F are schematic diagrams depicting various covalentlydimerized Gene Writer protein configurations. The proteins depicted are:FIG. 17A: a wild-type full length enzyme. FIG. 17B, two full-lengthenzymes (each comprising a DNA-binding domain, an RNA-binding domain, areverse transcriptase domain, and an endonuclease domain) connected by alinker. FIG. 17C, a DNA binding domain and an RNA binding domainconnected by a linker to a full-length enzyme. FIG. 17D, a DNA-bindingdomain and an RNA-binding domain connected by a linker to an RNA-bindingdomain, a reverse transcriptase domain, and an endonuclease domain. FIG.17E, a DNA-binding domain connected by a first linker to an RNA-bindingdomain, which is connected by a second linker to a second RNA-bindingdomain, a reverse transcriptase domain, and an endonuclease domain. FIG.17F, a DNA-binding domain connected by a first linker to an RNA-bindingdomain, which is connected by a second linker to a plurality ofRNA-binding domains (in this figure, the molecule comprises threeRNA-binding domains), which are connected by a linker to a reversetranscriptase domain and an endonuclease domain. In some embodiments,each R2 binds UTRs in the template RNA. In some embodiments, at leastone module comprises a reverse transcriptase domain and an endonucleasedomain. In some embodiments, the protein comprises a plurality ofRNA-binding domains. In some embodiments, the modular system is splitand is only active when it binds on DNA where the system uses twodifferent DNA binding modules, e.g., a first protein comprising a firstDNA binding module that is fused to an RNA binding module that recruitsthe RNA template for target primed reverse transcription, and secondprotein that comprises a second DNA binding module that binds at thesite of intergration and is fused to the reverse transcription andendonuclease modules. In some embodiments, the nucleic acid encoding theGeneWriter comprises an intein such that the GeneWriter protein isexpressed from two separate genes and is fused by protein splicing afterbeing translated. In some embodiments, the GeneWriter is derived from anon-LTR protein, e.g., an R2 protein.

FIGS. 18A-18F are a schematic diagram showing different modularcomponents of a GeneWriter protein. The proteins depicted are: FIG. 18A:a wild-type full length enzyme. FIG. 18B: the DNA-binding domain of aGeneWriter may comprise zinc fingers, Cas9, or a transcription factor,or a fragment or variant of any of the forgoing. FIG. 18C: the reversetranscriptase domain and RNA-binding domain together may comprise areverse transcriptase domain (e.g., from an R2 protein) that isheterologous to one or more other domains of the protein, and mayoptionally further comprise one or more additional RNA binding domains,or a fragment or variant of any of the foregoing. FIG. 18D: the RNAbinding domain may comprise, e.g., a B-box protein, an MS2 coat protein,a dCas protein, or a UTR binding protein, or a fragment or variant ofany of the foregoing. FIG. 18E: the reverse transcriptase domain maycomprise, e.g., a truncated reverse transcriptase domain, e.g., from anR2 protein; a reverse transcriptase domain from a virus (e.g., HIV), ora reverse transcriptase domain from AMV (avian myeloblastosis virus), ora fragment or variant of any of the foregoing. FIG. 18F: theendonuclease domain can comprise, e.g., a Cas9 nickase, a Cas ortholog,Fok I, or a restriction enzyme, or a fragment or variant of any of theforegoing. In some embodiments, a separate DNA binding domain can beattached to a polypeptide described herein (e.g., a DNA binding domainhaving stronger affinity for the target DNA sequence than an existing orprior DNA binding domain of the polypeptide, or a DNA binding sequencethat binds to a different target DNA sequence than the existing or priorDNA binding domain of the polypeptide). In some embodiments, DNA bindingdomain mutants can be generated, e.g., having increased affinity to thetarget DNA sequence. In embodiments, the DNA binding domain comprises azinc finger. In embodiments, the DNA binding domain is attached to thepolypeptide (e.g., at the N-terminal or C-terminal ends) via a linker,e.g., as described herein. In embodiments, a zinc finger is attached toa DNA binding domain mutant (e.g., as described herein), such that thepolypeptide exhibits increased binding to the target DNA sequence (e.g.,as dictated by the zinc finger) without competition with the rDNA.

FIG. 19 is a graph showing linker mutant integration into the genome ofHEK293T cells, assessed by a ddPCR assay evaluating copy number of R2Tgintegration per genome. In v1 mutants, an insertion is located at theN-terminal side of an alpha helical region of R2Tg that preceded thepredicted −1 RNA binding motif; in v2 mutants, an insertion is locatedat the C-terminal side of an alpha helical region of R2Tg that precededthe predicted −1 RNA binding motif; and in v3 mutants, an insertion islocated C-terminal to a random coil region that came after the predictedc-myb DNA binding motif of R2Tg.

FIGS. 20A-20B are a series of graphs showing long-read sequencingconfirming fidelity of R2Tg cis integration. Unique sequence coverage,as determined by UMI, is graphed across the expected reference sequence.The left vertical bar indicates expected 5′ junction of the rDNA andR2Tg, while the right vertical bar indicates the 3′ junction. Twoseparate amplicons spanning the 5′ junction and 3′ junction are shown.

FIGS. 21A-21B are a series of graphs showing long-read sequencingconfirming fidelity of R2Tg cis integration. Unique sequence deletions(>3 bp) as determined by UMI is graphed across the expected referencesequence. The left vertical bar indicates expected 5′ junction of therDNA and R2Tg, while the right vertical bar indicates the 3′ junction.Two separate amplicons spanning the 5′ junction and 3′ junction areshown.

FIG. 22 is a diagram showing exemplary plasmid map PLV033 for cisintegration of R2Gfo.

FIG. 23 is a graph showing integration of R2Gfo, R4A1, and R2Tg in cisin HEK293T cells. The mean of four replicates is shown; error barsindicate standard deviation.

FIG. 24 is a graph showing that R2Tg integrates into human fibroblastsin cis. Integration efficiency of the wild-type (WT) and endonuclease(EN) control R2Tg were plotted over four replicate experiments asmeasured via ddPCR at the 3′ junction of R2Tg and the rDNA target.

FIG. 25 is a diagram showing Western Blot analysis for p53, p21, Actin,and Vinculin. U2OS cells were tested with the indicated compound orplasmid: GFP, R2Tg-WT (wild-type), or R2Tg-EN (endonuclease domainmutant). Plasmid transfections were performed with either lipofectamine3000 (Lipo) or Fugene HD (Fug). Cells were analyzed 24 hours aftertreatment or transfection.

DETAILED DESCRIPTION

This disclosure relates to compositions, systems and methods fortargeting, editing, modifying or manipulating a DNA sequence (e.g.,inserting a heterologous object DNA sequence into a target site of amammalian genome) at one or more locations in a DNA sequence in a cell,tissue or subject, e.g., in vivo or in vitro. The object DNA sequencemay include, e.g., a coding sequence, a regulatory sequence, a geneexpression unit.

More specifically, the disclosure provides retrotransposon-based systemsfor inserting a sequence of interest into the genome. This disclosure isbased, in part, on a bioinformatic analysis to identify retrotransposasesequences and the associated 5′ UTR and 3′ UTR from a variety oforganisms (see Table 3). While not wishing to be bound by theory, insome embodiments, retrotransposases identified in homeothermic (warmblooded) species, like birds, may have improved thermostability relativeto some other enzymes that evolved at lower temperatures, and thethermostable retrotransposases may therefore be better suited for use inhuman cells. The disclosure also provides experimental evidence thatseveral retrotransposases from different species, e.g., differentspecies of animal and/or different species and clade of retrotransposon(e.g., as grouped by reverse transcriptase phylogeny, e.g., as describedin Su et al. (2019) RNA; incorporated herein by reference in itsentirety), can be used to catalyze DNA insertion into a target site inhuman cells (see Examples 7 and Example 28).

In some embodiments, systems described herein can have a number ofadvantages relative to various earlier systems. For instance, thedisclosure describes retrotransposases capable of inserting longsequences (e.g., over 3000 nucleotides) of heterologous nucleic acidinto a genome (see, e.g., FIG. 20A). In addition, retrotransposasesdescribed herein can insert heterologous nucleic acid in an endogenoussite in the genome, such as the rDNA locus (see, e.g., Example 7). Thisis in contrast to Cre/loxP systems which require a first step ofinserting an exogenous loxP site before a second step of inserting asequence of interest into the loxP site.

Gene-Writer™ Genome Editors

Non-long terminal repeat (LTR) retrotransposons are a type of mobilegenetic elements that are widespread in eukaryotic genomes. They includetwo classes: the apurinic/apyrimidinic endonuclease (APE)-type and therestriction enzyme-like endonuclease (RLE)-type. The APE classretrotransposons are comprised of two functional domains: anendonuclease/DNA binding domain, and a reverse transcriptase domain. TheRLE class are comprised of three functional domains: a DNA bindingdomain, a reverse transcription domain, and an endonuclease domain. Thereverse transcriptase domain of non-LTR retrotransposon functions bybinding an RNA sequence template and reverse transcribing it into thehost genome's target DNA. The RNA sequence template has a 3′untranslated region which is specifically bound to the transposase, anda variable 5′ region generally having Open Reading Frame(s) (“ORF”)encoding transposase proteins. The RNA sequence template may alsocomprise a 5′ untranslated region which specifically binds theretrotransposase.

The inventors have found that, surprisingly, the elements of suchnon-LTR retrotransposons can be functionally modularized and/or modifiedto target, edit, modify or manipulate a target DNA sequence, e.g., toinsert an object (e.g., heterologous) nucleic acid sequence into atarget genome, e.g., a mammalian genome, by reverse transcription. Suchmodularized and modified nucleic acids, polypeptide compositions andsystems are described herein and are referred to as Gene Writer™ geneeditors. A Gene Writer™ gene editor system comprises: (A) a polypeptideor a nucleic acid encoding a polypeptide, wherein the polypeptidecomprises (i) a reverse transcriptase domain, and either (x) anendonuclease domain that contains DNA binding functionality or (y) anendonuclease domain and separate DNA binding domain; and (B) a templateRNA comprising (i) a sequence that binds the polypeptide and (ii) aheterologous insert sequence. For example, the Gene Writer genome editorprotein may comprise a DNA-binding domain, a reverse transcriptasedomain, and an endonuclease domain. In other embodiments, the GeneWriter genome editor protein may comprise a reverse transcriptase domainand an endonuclease domain. In certain embodiments, the elements of theGene Writer™ gene editor polypeptide can be derived from sequences ofnon-LTR retrotransposons, e.g., APE-type or RLE-type retrotransposons orportions or domains thereof. In some embodiments the RLE-type non-LTRretrotransposon is from the R2, NeSL, HERO, R4, or CRE clade. In someembodiments the Gene Writer genome editor is derived from R4 elementX4_Line, which is found in the human genome. In some embodiments theAPE-type non-LTR retrotransposon is from the R1, or Tx1 clade. In someembodiments the Gene Writer genome editor is derived from Tx1 elementMare6, which is found in the human genome. The RNA template element of aGene Writer™ gene editor system is typically heterologous to thepolypeptide element and provides an object sequence to be inserted(reverse transcribed) into the host genome. In some embodiments the GeneWriter genome editor protein is capable of target primed reversetranscription.

In some embodiments the Gene Writer genome editor is combined with asecond polypeptide. In some embodiments the second polypeptide isderived from an APE-type non-LTR retrotransposon. In some embodimentsthe second polypeptide has a zinc knuckle-like motif. In someembodiments the second polypeptide is a homolog of Gag proteins.

Polypeptide Component of Gene Writer Gene Editor System

RT Domain:

In certain aspects of the present invention, the reverse transcriptasedomain of the Gene Writer system is based on a reverse transcriptasedomain of an APE-type or RLE-type non-LTR retrotransposon. A wild-typereverse transcriptase domain of an APE-type or RLE-type non-LTRretrotransposon can be used in a Gene Writer system or can be modified(e.g., by insertion, deletion, or substitution of one or more residues)to alter the reverse transcriptase activity for target DNA sequences. Insome embodiments the reverse transcriptase is altered from its naturalsequence to have altered codon usage, e.g. improved for human cells. Insome embodiments the reverse transcriptase domain is a heterologousreverse transcriptase from a different retrovirus, LTR-retrotransposon,or non-LTR retrotransposon. In certain embodiments, a Gene Writer systemincludes a polypeptide that comprises a reverse transcriptase domain ofan RLE-type non-LTR retrotransposon from the R2, NeSL, HERO, R4, or CREclade, or of an APE-type non-LTR retrotransposon from the R1, or Tx1clade. In certain embodiments, a Gene Writer system includes apolypeptide that comprises a reverse transcriptase domain of aretrotransposon listed in Table 1, Table 2, or Table 3. In embodiments,the amino acid sequence of the reverse transcriptase domain of a GeneWriter system is at least about 50%, at least about 60%, at least about70%, at least about 80%, at least about 85%, at least about 90%, atleast about 95%, at least about 96%, at least about 97%, at least about98%, at least about 99% identical to the amino acid sequence of areverse transcriptase domain of a retrotransposon whose DNA sequence isreferenced in Table 1, Table 2, or Table 3. A person having ordinaryskill in the art is capable of identifying reverse transcription domainsbased upon homology to other known reverse transcription domains usingroutine tools as Basic Local Alignment Search Tool (BLAST). In someembodiments, reverse transcriptase domains are modified, for example bysite-specific mutation. In embodiments, the reverse transcriptase domainis engineered to bind a heterologous template RNA.

Endonuclease Domain:

In certain embodiments, the endonuclease/DNA binding domain of anAPE-type retrotransposon or the endonuclease domain of an RLE-typeretrotransposon can be used or can be modified (e.g., by insertion,deletion, or substitution of one or more residues) in a Gene Writersystem described herein. In some embodiments the endonuclease domain orendonuclease/DNA binding domain is altered from its natural sequence tohave altered codon usage, e.g. improved for human cells. In someembodiments the endonuclease element is a heterologous endonucleaseelement, such as Fok1 nuclease, a type-II restriction 1-likeendonuclease (RLE-type nuclease), or another RLE-type endonuclease (alsoknown as REL). In some embodiments the heterologous endonucleaseactivity has nickase activity and does not form double stranded breaks.The amino acid sequence of an endonuclease domain of a Gene Writersystem described herein may be at least about 50%, at least about 60%,at least about 70%, at least about 80%, at least about 85%, at leastabout 90%, at least about 95%, at least about 96%, at least about 97%,at least about 98%, at least about 99% identical to the amino acidsequence of an endonuclease domain of a retrotransposon whose DNAsequence is referenced in Table 1, 2, or 3. A person having ordinaryskill in the art is capable of identifying endonuclease domains basedupon homology to other known endonuclease domains using tools as BasicLocal Alignment Search Tool (BLAST). In certain embodiments, theheterologous endonuclease is Fok1 or a functional fragment thereof. Incertain embodiments, the heterologous endonuclease is a Hollidayjunction resolvase or homolog thereof, such as the Holliday junctionresolving enzyme from Sulfolobus solfataricus—Ssol Hje (Govindaraju etal., Nucleic Acids Research 44:7, 2016). In certain embodiments, theheterologous endonuclease is the endonuclease of the large fragment of aspliceosomal protein, such as Prp8 (Mahbub et al., Mobile DNA 8:16,2017). For example, a Gene Writer polypeptide described herein maycomprise a reverse transcriptase domain from an APE- or RLE-typeretrotransposon and an endonuclease domain that comprises Fok1 or afunctional fragment thereof. In still other embodiments, homologousendonuclease domains are modified, for example by site-specificmutation, to alter DNA endonuclease activity. In still otherembodiments, endonuclease domains are modified to remove any latentDNA-sequence specificity.

DNA Binding Domain:

In certain aspects, the DNA-binding domain of a Gene Writer polypeptidedescribed herein is selected, designed, or constructed for binding to adesired host DNA target sequence. In certain embodiments, theDNA-binding domain of the engineered RLE is a heterologous DNA-bindingprotein or domain relative to a native retrotransposon sequence. In someembodiments the heterologous DNA binding element is a zinc-fingerelement or a TAL effector element, e.g., a zinc-finger or TALpolypeptide or functional fragment thereof. In some embodiments theheterologous DNA binding element is a sequence-guided DNA bindingelement, such as Cas9, Cpf1, or other CRISPR-related protein that hasbeen altered to have no endonuclease activity. In some embodiments theheterologous DNA binding element retains endonuclease activity. In someembodiments the heterologous DNA binding element replaces theendonuclease element of the polypeptide. In specific embodiments, theheterologous DNA-binding domain can be any one or more of Cas9, TALdomain, ZF domain, Myb domain, combinations thereof, or multiplesthereof. In certain embodiments, the heterologous DNA-binding domain isa DNA binding domain of a retrotransposon described in Table 1, Table 2,or Table 3. A person having ordinary skill in the art is capable ofidentifying DNA binding domains based upon homology to other known DNAbinding domains using tools as Basic Local Alignment Search Tool(BLAST). In still other embodiments, DNA-binding domains are modified,for example by site-specific mutation, increasing or decreasingDNA-binding elements (for example, number and/or specificity of zincfingers), etc., to alter DNA-binding specificity and affinity. In someembodiments the DNA binding domain is altered from its natural sequenceto have altered codon usage, e.g. improved for human cells

In certain aspects of the present invention, the host DNA-binding siteintegrated into by the Gene Writer system can be in a gene, in anintron, in an exon, an ORF, outside of a coding region of any gene, in aregulatory region of a gene, or outside of a regulatory region of agene. In other aspects, the engineered RLE may bind to one or more thanone host DNA sequence.

In certain embodiments, a Gene Writer™ gene editor system RNA furthercomprises an intracellular localization sequence, e.g., a nuclearlocalization sequence. The nuclear localization sequence may be an RNAsequence that promotes the import of the RNA into the nucleus. Incertain embodiments the nuclear localization signal is located on thetemplate RNA. In certain embodiments, the retrotransposase polypeptideis encoded on a first RNA, and the template RNA is a second, separate,RNA, and the nuclear localization signal is located on the template RNAand not on an RNA encoding the retrotransposase polypeptide. While notwishing to be bound by theory, in some embodiments, the RNA encoding theretrotransposase is targeted primarily to the cytoplasm to promote itstranslation, while the template RNA is targeted primarily to the nucleusto promote its retrotransposition into the genome. In some embodimentsthe nuclear localization signal is at the 3′ end, 5′ end, or in aninternal region of the template RNA. In some embodiments the nuclearlocalization signal is 3′ of the heterologous sequence (e.g., isdirectly 3′ of the heterologous sequence) or is 5′ of the heterologoussequence (e.g., is directly 5′ of the heterologous sequence). In someembodiments the nuclear localization signal is placed outside of the 5′UTR or outside of the 3′ UTR of the template RNA. In some embodimentsthe nuclear localization signal is placed between the 5′ UTR and the 3′UTR, wherein optionally the nuclear localization signal is nottranscribed with the transgene (e.g., the nuclear localization signal isan anti-sense orientation or is downstream of a transcriptionaltermination signal or polyadenylation signal). In some embodiments thenuclear localization sequence is situated inside of an intron. In someembodiments a plurality of the same or different nuclear localizationsignals are in the RNA, e.g., in the template RNA. In some embodimentsthe nuclear localization signal is less than 5, 10, 25, 50, 75, 100,150, 200, 250, 300, 350, 400, 450, 500, 600, 700, 800, 900 or 1000 bp inlength. Various RNA nuclear localization sequences can be used. Forexample, Lubelsky and Ulitsky, Nature 555 (107-111), 2018 describe RNAsequences which drive RNA localization into the nucleus. In someembodiments, the nuclear localization signal is a SINE-derived nuclearRNA localization (SIRLOIN) signal. In some embodiments the nuclearlocalization signal binds a nuclear-enriched protein. In someembodiments the nuclear localization signal binds the HNRNPK protein. Insome embodiments the nuclear localization signal is rich in pyrimidines,e.g., is a C/T rich, C/U rich, C rich, T rich, or U rich region. In someembodiments the nuclear localization signal is derived from a longnon-coding RNA. In some embodiments the nuclear localization signal isderived from MALAT1 long non-coding RNA or is the 600 nucleotide Mregion of MALAT1 (described in Miyagawa et al., RNA 18, (738-751),2012). In some embodiments the nuclear localization signal is derivedfrom BORG long non-coding RNA or is a AGCCC motif (described in Zhang etal., Molecular and Cellular Biology 34, 2318-2329 (2014). In someembodiments the nuclear localization sequence is described in Shukla etal., The EMBO Journal e98452 (2018). In some embodiments the nuclearlocalization signal is derived from a non-LTR retrotransposon, an LTRretrotransposon, retrovirus, or an endogenous retrovirus.

In certain embodiments, a Gene Writer™ gene editor system polypeptidefurther comprises an intracellular localization sequence, e.g., anuclear localization sequence and/or a nucleolar localization sequence.The nuclear localization sequence and/or nucleolar localization sequencemay be amino acid sequences that promote the import of the protein intothe nucleus and/or nucleolus, where it can promote integration ofheterologous sequence into the genome. In certain embodiments, a GeneWriter gene editor system polypeptide (e.g., a retrotransposase, e.g., apolypeptide according to any of Tables 1, 2, or 3 herein) furthercomprises a nucleolar localization sequence. In certain embodiments, theretrotransposase polypeptide is encoded on a first RNA, and the templateRNA is a second, separate, RNA, and the nucleolar localization signal isencoded on the RNA encoding the retrotransposase polypeptide and not onthe template RNA. In some embodiments, the nucleolar localization signalis located at the N-terminus, C-terminus, or in an internal region ofthe polypeptide. In some embodiments, a plurality of the same ordifferent nucleolar localization signals are used. In some embodiments,the nuclear localization signal is less than 5, 10, 25, 50, 75, or 100amino acids in length. Various polypeptide nucleolar localizationsignals can be used. For example, Yang et al., Journal of BiomedicalScience 22, 33 (2015), describe a nuclear localization signal that alsofunctions as a nucleolar localization signal. In some embodiments, thenucleolar localization signal may also be a nuclear localization signal.In some embodiments, the nucleolar localization signal may overlap witha nuclear localization signal. In some embodiments, the nucleolarlocalization signal may comprise a stretch of basic residues. In someembodiments, the nucleolar localization signal may be rich in arginineand lysine residues. In some embodiments, the nucleolar localizationsignal may be derived from a protein that is enriched in the nucleolus.In some embodiments, the nucleolar localization signal may be derivedfrom a protein enriched at ribosomal RNA loci. In some embodiments, thenucleolar localization signal may be derived from a protein that bindsrRNA. In some embodiments, the nucleolar localization signal may bederived from MSP58. In some embodiments, the nucleolar localizationsignal may be a monopartite motif. In some embodiments, the nucleolarlocalization signal may be a bipartite motif. In some embodiments, thenucleolar localization signal may consist of a multiple monopartite orbipartite motifs. In some embodiments, the nucleolar localization signalmay consist of a mix of monopartite and bipartite motifs. In someembodiments, the nucleolar localization signal may be a dual bipartitemotif. In some embodiments, the nucleolar localization motif may be aKRASSQALGTIPKRRSSSRFIKRKK (SEQ ID NO: 1530). In some embodiments, thenucleolar localization signal may be derived from nuclearfactor-KB-inducing kinase. In some embodiments, the nucleolarlocalization signal may be an RKKRKKK motif (SEQ ID NO: 1531) (describedin Birbach et al., Journal of Cell Science, 117 (3615-3624), 2004).

In some embodiments, a nucleic acid described herein (e.g., an RNAencoding a GeneWriter polypeptide, or a DNA encoding the RNA) comprisesa microRNA binding site. In some embodiments, the microRNA binding siteis used to increase the target-cell specificity of a GeneWriter system.For instance, the microRNA binding site can be chosen on the basis thatis is recognized by a miRNA that is present in a non-target cell type,but that is not present (or is present at a reduced level relative tothe non-target cell) in a target cell type. Thus, when the RNA encodingthe GeneWriter polypeptide is present in a non-target cell, it would bebound by the miRNA, and when the RNA encoding the GeneWriter polypeptideis present in a target cell, it would not be bound by the miRNA (orbound but at reduced levels relative to the non-target cell). While notwishing to be bound by theory, binding of the miRNA to the RNA encodingthe GeneWriter polypeptide may reduce production of the GeneWriterpolypeptide, e.g., by degrading the mRNA encoding the polypeptide or byinterfering with translation. Accordingly, the heterologous objectsequence would be inserted into the genome of target cells moreefficiently than into the genome of non-target cells. A system having amicroRNA binding site in the RNA encoding the GeneWriter polypeptide (orencoded in the DNA encoding the RNA) may also be used in combinationwith a template RNA that is regulated by a second microRNA binding site,e.g., as described herein in the section entitled “Template RNAcomponent of Gene Writer™ gene editor system.”

TABLE 1 Table 1: APE-type non-LTR retrotransposon elements SequenceFamily Accession Mobile Element Name Organism Dewa AB097143 ORF2 Daniorerio retrotransposon Danio rerio DewaDr1 DNA, complete sequence HeT-AKJ081250 non-LTR Drosophila melanogaster non-LTR Drosophilaretrotransposon: retrotransposon HeT-A, partial melanogaster HeT-Asequence Keno AB111948 ORF2 Tetraodon nigroviridis Tetraodonretrotransposon KenoTn1 DNA, nigroviridis partial sequence KenoDr1;AB097144 ORF2 Danio rerio retrotransposon KenoDr1 Danio rerio Keno DNA,complete sequence KenoFr1; AB111947 ORF2 Takifugu rubripesretrotransposon Takifugu Keno KenoFr1 DNA, complete sequence rubripesKibi AB097139 ORF2 Danio rerio retrotransposon KibiDr2 Danio rerio DNA,complete sequence Kibi AB097138 ORF2 Danio rerio retrotransposon KibiDr1Danio rerio DNA, complete sequence Kibi AB097137 ORF2 Tetraodonnigroviridis Tetraodon retrotransposon KibiTn1 DNA, nigroviridiscomplete sequence Kibi AB097136 ORF2 Takifugu rubripes retrotransposonTakifugu KibiFr1 DNA, complete sequence rubripes KoshiTn1 AB097135 ORF2Tetraodon nigroviridis Tetraodon retrotransposon KoshiTn1 DNA,nigroviridis complete sequence Mutsu AB097142 ORF2 Danio rerioretrotransposon Danio rerio MutsuDr3 DNA, partial sequence MutsuAB097141 ORF2 Danio rerio retrotransposon Danio rerio MutsuDr2 DNA,partial sequence Mutsu AB097140 ORF2 Danio rerio retrotransposon Daniorerio MutsuDr1 DNA, complete sequence R1 HQ284568 non-LTR Trilocha sp.GAS-2011 isolate TrilSp.6 Trilocha retrotransposon: non-LTRretrotransposon R1-like sp. GAS-2011 R1-like reverse transcriptase gene,partial cds R1 HQ284534 non-LTR Scopula ornata isolate ScoOrn.6 non-Scopula ornata retrotransposon: LTR retrotransposon R1-like reverseR1-like transcriptase gene, partial cds R1 HQ284496 non-LTR Perigoniailus isolate PerIlus.31 non- Perigonia ilus retrotransposon: LTRretrotransposon R1-like reverse R1-like transcriptase gene, partial cdsR1 HQ284489 non-LTR Oxytenis modestia isolate Oxytenis retrotransposon:OxyMod.2_3_4_7_9 non-LTR modestia R1-like retrotransposon R1-likereverse transcriptase gene, partial cds R1 HQ284488 non-LTR Oxytenismodestia isolate OxyMod.1 Oxytenis retrotransposon: non-LTRretrotransposon R1-like modestia R1-like reverse transcriptase gene,partial cds R1 HQ284476 non-LTR Oeneis magna dubia isolate Oeneis magnaretrotransposon: OenMag.26 non-LTR retrotransposon dubia R1-like R1-likereverse transcriptase-like gene, partial sequence R1 HQ284437 non-LTRLymantria dispar isolate LymDis.2 Lymantria retrotransposon: non-LTRretrotransposon R1-like dispar R1-like reverse transcriptase gene,partial cds R1 HQ284435 non-LTR Lymantria dispar isolate LymDis.1Lymantria retrotransposon: non-LTR retrotransposon R1-like disparR1-like reverse transcriptase gene, partial cds R1 HQ284432 non-LTRJaniodes laverna isolate JanLav.911 Janiodes retrotransposon: non-LTRretrotransposon R1-like laverna R1-like reverse transcriptase-like gene,partial sequence R1 HQ284431 non-LTR Janiodes laverna isolate JanLav.811Janiodes retrotransposon: non-LTR retrotransposon R1-like lavernaR1-like reverse transcriptase gene, partial cds R1 HQ284430 non-LTRJaniodes laverna isolate JanLav.5 Janiodes retrotransposon: non-LTRretrotransposon R1-like laverna R1-like reverse transcriptase gene,partial cds R1 HQ284428 non-LTR Janiodes laverna isolate JanLav.411Janiodes retrotransposon: non-LTR retrotransposon R1-like lavernaR1-like reverse transcriptase gene, partial cds R1 HQ284426 non-LTRJaniodes laverna isolate JanLav.211 Janiodes retrotransposon: non-LTRretrotransposon R1-like laverna R1-like reverse transcriptase-like gene,partial sequence R1 HQ284421 non-LTR Heteropterus morpheus isolateHeteropterus retrotransposon: HetMor.3 non-LTR retrotransposon morpheusR1-like R1-like reverse transcriptase gene, partial cds R1 HQ284402non-LTR Erinnyis ello isolate EriEllo.22 non- Erinnyis elloretrotransposon: LTR retrotransposon R1-like reverse R1-liketranscriptase-like gene, partial sequence R1 HQ284399 non-LTR Erebiatheano isolate EreThe.29 non- Erebia theano retrotransposon: LTRretrotransposon R1-like reverse R1-like transcriptase gene, partial cdsR1 HQ284398 non-LTR Erebia theano isolate EreThe.28 non- Erebia theanoretrotransposon: LTR retrotransposon R1-like reverse R1-liketranscriptase-like gene, partial sequence R1 HQ284397 non-LTR Erebiatheano isolate EreThe.27 non- Erebia theano retrotransposon: LTRretrotransposon R1-like reverse R1-like transcriptase-like gene, partialsequence R1 HQ284391 non-LTR Emesis lucinda isolate EmeLuc.23 Emesislucinda retrotransposon: non-LTR retrotransposon R1-like R1-like reversetranscriptase gene, partial cds R1 HQ284390 non-LTR Emesis lucindaisolate EmeLuc.2 non- Emesis lucinda retrotransposon: LTRretrotransposon R1-like reverse R1-like transcriptase gene, partial cdsR1 HQ284364 non-LTR Coenonympha glycerion isolate Coenonympharetrotransposon: CoeGly.9 non-LTR retrotransposon glycerion R1-likeR1-like reverse transcriptase gene, partial cds R1 HQ284363 non-LTRCoenonympha glycerion isolate Coenonympha retrotransposon: CoeGly.8non-LTR retrotransposon glycerion R1-like R1-like reversetranscriptase-like gene, partial sequence R1 HQ284362 non-LTRCoenonympha glycerion isolate Coenonympha retrotransposon: CoeGly.7non-LTR retrotransposon glycerion R1-like R1-like reverse transcriptasegene, partial cds R1 HQ284361 non-LTR Coenonympha glycerion isolateCoenonympha retrotransposon: CoeGly.5 non-LTR retrotransposon glycerionR1-like R1-like reverse transcriptase-like gene, partial sequence R1HQ284357 non-LTR Coenonympha glycerion isolate Coenonympharetrotransposon: CoeGly.13 non-LTR retrotransposon glycerion R1-likeR1-like reverse transcriptase gene, partial cds R1 HQ284356 non-LTRCoenonympha glycerion isolate Coenonympha retrotransposon: CoeGly.11non-LTR retrotransposon glycerion R1-like R1-like reversetranscriptase-like gene, partial sequence R1 HQ284350 non-LTRCatocyclotis adelina isolate Catocyclotis retrotransposon: CatAde.18non-LTR retrotransposon adelina R1-like R1-like reverse transcriptasegene, partial cds R1 HQ284340 non-LTR Caria rhacotis isolate CarRha.11non- Caria rhacotis retrotransposon: LTR retrotransposon R1-like reverseR1-like transcriptase gene, partial cds R1 HQ284339 non-LTR Cariarhacotis isolate CarRha.1 non- Caria rhacotis retrotransposon: LTRretrotransposon R1-like reverse R1-like transcriptase gene, partial cdsR1 HQ284319 non-LTR Archiearis parthenias isolate BrePar.1 Archiearisretrotransposon: non-LTR retrotransposon R1-like parthenias R1-likereverse transcriptase gene, partial cds R1 HQ284318 non-LTR Brangasneora isolate BraNeo.32 Brangas neora retrotransposon: non-LTRretrotransposon R1-like R1-like reverse transcriptase gene, partial cdsR1 HQ284292 non-LTR Araschnia levana isolate AraLev.31 Araschnia levanaretrotransposon: non-LTR retrotransposon R1-like R1-like reversetranscriptase-like gene, partial sequence R1 HQ284286 non-LTR Araschnialevana isolate AraLev.1 Araschnia levana retrotransposon: non-LTRretrotransposon R1-like R1-like reverse transcriptase gene, partial cdsR1 HQ284280 non-LTR Anteros formosus isolate AntForm.34 Anterosretrotransposon: non-LTR retrotransposon R1-like formosus R1-likereverse transcriptase gene, partial cds R1 HQ284279 non-LTR Anterosformosus isolate AntForm.32 Anteros retrotransposon: non-LTRretrotransposon R1-like formosus R1-like reverse transcriptase-likegene, partial sequence R1 HQ284278 non-LTR Anteros formosus isolateAntForm.31 Anteros retrotransposon: non-LTR retrotransposon R1-likeformosus R1-like reverse transcriptase-like gene, partial sequence R1HQ284270 non-LTR Agrotis exclamationis isolate Agrotis retrotransposon:AgrExcl.27 non-LTR retrotransposon exclamationis R1-like R1-like reversetranscriptase gene, partial cds R1 HQ284267 non-LTR Agrius cingulataisolate Agrius retrotransposon: AgrCing.36_39 non-LTR cingulata R1-likeretrotransposon R1-like reverse transcriptase gene, partial cds R1HQ284266 non-LTR Agrius cingulata isolate AgrCing.3 Agriusretrotransposon: non-LTR retrotransposon R1-like cingulata R1-likereverse transcriptase-like gene, partial sequence R1 HQ284263 non-LTRAglia tau isolate AglTau.8 non-LTR Aglia tau retrotransposon:retrotransposon R1-like reverse R1-like transcriptase gene, partial cdsR1 HQ284262 non-LTR Aglia tau isolate AglTau.7 non-LTR Aglia tauretrotransposon: retrotransposon R1-like reverse R1-like transcriptasegene, partial cds R1 DQ836362 MalR1 Maculinea alcon R1-like non-LTRPhengaris alcon retrotransposon R1 reverse transcriptase (RT)pseudogene, partial sequence R1 DQ836391 MnaR1 Maculinea nausithousR1-like non- Phengaris LTR retrotransposon R1 reverse nausithoustranscriptase (RT) gene, partial cds R1 KU543683 non-LTR Bactroceratryoni clone Btry_5404 Bactrocera retrotransposon non-LTRretrotransposon R1, tryoni and non-LTR complete sequence retrovirusreverse transcriptase; Region: RT_nLTR R1 KU543682 non-LTR Bactroceratryoni clone Btry_5167 Bactrocera retrotransposon non-LTRretrotransposon R1, tryoni and non-LTR complete sequence retrovirusreverse transcriptase; Region: RT_nLTR R1 KU543679 non-LTR Bactroceratryoni clone Btry_4956 Bactrocera retrotransposon non-LTRretrotransposon R1, tryoni and non-LTR complete sequence retrovirusreverse transcriptase; Region: RT_nLTR R1 KU543678 non-LTR Bactroceratryoni clone Btry_5979 Bactrocera retrotransposon non-LTRretrotransposon R1, tryoni and non-LTR complete sequence retrovirusreverse transcriptase; Region: RT_nLTR R1 AB078933 ORF1 Papilio xuthusnon-LTR Papilio xuthus retrotransposon gene for gag-like protein,partial cds, clone: SARTPx2-2 R1 AB078932 ORF1 Papilio xuthus non-LTRPapilio xuthus retrotransposon gene for gag-like protein, partial cds,clone: SARTPx2-1 R1 AB078936 ORF2 Papilio xuthus non-LTR Papilio xuthusretrotransposon genes for gag-like protein, reverse transcriptase,partial cds, clone: SARTPx4-N18 R1 AB078935 ORF2 Papilio xuthus non-LTRPapilio xuthus retrotransposon genes for gag-like protein, reversetranscriptase, partial and complete cds, clone: SARTPx3-N7 R1 AB078934ORF2 Papilio xuthus non-LTR Papilio xuthus retrotransposon genes forgag-like protein, reverse transcriptase, partial cds, clone: SARTPx3-N3R1 AB078931 ORF2 Papilio xuthus non-LTR Papilio xuthus retrotransposongenes for gag-like protein, reverse transcriptase, partial and completecds, clone: SARTPx1-N14 R1 AB078930 ORF2 Papilio xuthus non-LTR Papilioxuthus retrotransposon genes for gag-like protein, reversetranscriptase, partial and complete cds, clone: SARTPx1-N5 R1 AB078929ORF2 Papilio xuthus non-LTR Papilio xuthus retrotransposon genes forgag-like protein, reverse transcrpitase, partial and complete cds,clone: SARTPx1-N4 R1 AB078928 ORF2 Papilio xuthus non-LTR Papilio xuthusretrotransposon gene for gag-like protein, reverse transcrpitase,complete and partial cds, clone: SARTPx1-3 R1 KP771712 ORF2; containsBlattella germanica non-LTR Blattella endonuclease, retrotransposonTRAS-like 2, germanica reverse complete sequence transcriptase andRNaseH R1 KP771711 ORF2; contains Blattella germanica non-LTR Blattellaendonuclease, retrotransposon TRAS-like 1, germanica reverse completesequence transcriptase and RNaseH R1 AF015813 R1 ORF Dugesiella sp.retrotransposon R1 Aphonopelma reverse transcriptase gene, partial sp.WDB-1998 cds R1 AF015489 R1 ORF Dugesiella sp. retrotransposon R1Aphonopelma reverse transcriptase gene, partial sp. WDB-1998 cds R1BmAB182560 non-LTR Bombyx mori genes for non-LTR Bombyx moriretrotransposon retrotransposon R1Bmks ORF1 R1Bmks ORF2 protein, non-LTRretrotransposon R1Bmks ORF2 protein, complete cds R6 AB090819 ORF2Anopheles gambiae retrotransposon Anopheles R6Ag3 DNA, complete sequencegambiae R6 AB090818 ORF2 Anopheles gambiae retrotransposon AnophelesR6Ag2 DNA, complete sequence gambiae R6 AB090817 ORF2 Anopheles gambiaeretrotransposon Anopheles R6Ag1 DNA, complete sequence gambiae R6KJ958615 R2 Bacillus rossius non-LTR Bacillus retrotransposon reVIR6,partial rossius sequence R6 KJ958596 R2 Bacillus rossius non-LTRBacillus retrotransposon reBER6, partial rossius sequence R6 AF352480transposon: Chironomus circumdatus clone cir6 Chironomus NLRCth1-transposon NLRCth1-like non-LTR circumdatus like non-LTR retrotransposonreverse retrotransposon transcriptase gene, partial cds R6 AF373367transposon: Clelia rustica clone CR6 non-LTR Paraphimophis non-LTRretrotransposon LINE2 reverse rusticus retrotransposon transcriptasepseudogene, partial LINE2 sequence R7 AB090820 ORF2 Anopheles gambiaeretrotransposon Anopheles R7Ag1 DNA, complete sequence gambiae R7AB090821 ORF2 Anopheles gambiae retrotransposon Anopheles R7Ag2 DNA,complete sequence gambiae R7 KJ958622 R2 Bacillus rossius non-LTRBacillus retrotransposon trKOR7, partial rossius sequence R7 KJ958616 R2Bacillus rossius non-LTR Bacillus retrotransposon reVIR7, partialrossius sequence R7 KJ958597 R2 Bacillus rossius non-LTR Bacillusretrotransposon reBER7, partial rossius sequence R7 AF352514 transposon:Chironomus circumdatus clone cir7 Chironomus NLRCth1-like transposonNLRCth1-like non-LTR circumdatus non-LTR retrotransposon reverseretrotransposon transcriptase pseudogene, partial sequence Rt2 AY379084truncated; Leptocheirus plumulosus Leptocheirus similar toretrotransposon LpRt2, partial plumulosus reverse sequence transcriptaseRt2 MSQRT2RET Anopheles gambiae retrotransposon Anopheles RT2, completesequence gambiae RTAg4 AB090813 ORF2 Anopheles gambiae retrotransposonAnopheles RTAg4 DNA, complete sequence gambiae TRAS1 BMOTRAS1 DNAbinding Bombyx mori gene, complete Bombyx mori domain at sequence ofretrotransposon TRAS1 AA1103-1120. TRAS3 JX875955 similar toAcyrthosiphon pisum clone LSR1 non- Acyrthosiphon reverse LTRretrotransposon TRAS3, pisum transcriptases complete sequence Tx1AJ621359 transposon: Tetraodon nigroviridis non-LTR Tetraodon non-LTRretrotransposon TX1-1_Tet, complete nigroviridis retrotransposonsequence TX1-1_Tet Tx1 AJ621360 transposon: Tetraodon nigroviridispartial non- Tetraodon non-LTR LTR retrotransposon TX1-2_Tetnigroviridis retrotransposon TX1-2_Tet Tx1 AJ621361 transposon:Tetraodon nigroviridis partial non- Tetraodon non-LTR LTRretrotransposon TX1-3_Tet nigroviridis retrotransposon TX1-3_Tet Tx1AJ621362 transposon: Tetraodon nigroviridis partial non- Tetraodonnon-LTR LTR retrotransposon TX1-4_Tet nigroviridis retrotransposonTX1-4_Tet Tx1 DQ118004 transposon: Acipenser ruthenus clone dg194Acipenser Tx1-like transposon Tx1-like retrotransposon ruthenusretrotransposon Tx1Aru reverse transcriptase-like Tx1Aru gene, partialsequence Tx1 AB097134 ORF2 Takifugu rubripes retrotransposon TakifuguKoshiFr1 DNA, complete sequence rubripes Tx1 AB090816 ORF2 Anophelesgambiae retrotransposon Anopheles MinoAg1 DNA, complete sequence gambiaeTx1 AB090812 ORF2 Anopheles gambiae retrotransposon Anopheles RTAg3 DNA,complete sequence gambiae Waldo AH009917 non-LTR Drosophila melanogasterWaldo-A Drosophila retrotransposon: non-LTR retrotransposon, 5′melanogaster Waldo-A sequence Waldo AH009916 non-LTR Drosophilamelanogaster clone CBE9 Drosophila retrotransposon: Waldo-A non-LTRretrotransposon, 5′ melanogaster Waldo-A sequence Waldo AH009915 non-LTRDrosophila melanogaster Waldo-A Drosophila retrotransposon: non-LTRretrotransposon, 5′ melanogaster Waldo-A sequence Waldo AH009914 non-LTRDrosophila melanogaster Waldo-A Drosophila retrotransposon: non-LTRretrotransposon melanogaster Waldo-A Waldo AH009920 non-LTR Drosophilamelanogaster Waldo-B Drosophila retrotransposon: non-LTRretrotransposon, 5′ melanogaster Waldo-B sequence Waldo AH009919 non-LTRDrosophila retrotransposon: melanogaster Waldo-B Waldo AH009918 non-LTRDrosophila retrotransposon: melanogaster Waldo-B Waldo AB090815 ORF2Anopheles gambiae retrotransposon Anopheles WaldoAg2 DNA, completesequence gambiae Waldo AB090814 ORF2 Anopheles gambiae retrotransposonAnopheles WaldoAg1 DNA, complete sequence gambiae Waldo AB078939 ORF2Forficula scudderi non-LTR Forficula retrotransposon pseudogene forscudderi reverse transcriptase, clone: WaldoFs1-26 Waldo AB078938 ORF2Forficula scudderi non-LTR Forficula retrotransposon pseudogene forscudderi reverse transcriptase, clone: WaldoFs1-2 Waldo AB078937 ORF2Forficula scudderi non-LTR Forficula retrotransposon pseudogene forscudderi reverse transcriptase, clone: WaldoFs1-1

TABLE 2 Table 2: RLE-type non-LTR retrotransposon elements FamilyAccession Mobile Element Name/Description Organism CRE EF067892Colletotrichum cereale Colletotrichum clone 9F8-1558 Ccret3 non-LTRcereale retrotransposon, partial sequence CRE EF067894 Colletotrichumcereale Colletotrichum clone 9F8-2137 Ccret3 non-LTR cerealeretrotransposon, partial sequence CRE MG028000 non-LTR Characidiumgomesi voucher Characidium retrotransposon: MNRJ20998 non-LTR gomesiRex3 retrotransposon Rex3, partial sequence CRE KY566213 non-LTRCharacidium gomesi non-LTR Characidim retrotransposon: retrotransposonRex3, partial gomesi Rex3 sequence CRE GU949558 Kalotermes flavicollisKalotermes isolate Crete non-LTR flavicollis retrotransposon R2,complete sequence; and R2 protein gene, complete cds CRE; CFU19151 polydA Crithidia fasciculata Crithidia CRE2 tracts in retrotransposon CRE2in mini- fasciculata 5′ and exon gene, putative reverse 3′ UTRstranscriptase gene, complete cds CZAR BR000987 pol TPA_inf: Capsasporaowczarzaki Capsaspora DNA, non-LTR retrotransposon owczarzaki CoL4,complete sequence, strain: ATCC 30864 CZAR BR000986 pol TPA_inf:Capsaspora owczarzaki Capsaspora DNA, non-LTR retrotransposon owczarzakiCoL3, complete sequence, strain: ATCC 30864 CZAR BR000985 pol TPA_inf:Capsaspora owczarzaki Capsaspora DNA, non-LTR retrotransposon owczarzakiCoL2, complete sequence, strain: ATCC 30864 CZAR BR000984 pol TPA_inf:Capsaspora owczarzaki Capsaspora DNA, non-LTR retrotransposon owczarzakiCoL1, complete sequence, strain: ATCC 30864 DongAG; AB097127 rtAnopheles gambiae Anopheles Dong retrotransposon DongAg DNA, gambiaepartial sequence EhRLE2 AB097128 rt Entamoeba histolytica Entamoebaretrotransposon EhRLE2 DNA, histolytica complete sequence EhRLE3AB097129 rt Entamoeba histolytica Entamoeba retrotransposon EhRLE3 DNA,histolytica complete sequence Genie AF440196 endonuclease Giardiaintestinalis non-LTR Giardia retrotransposon GENIE 1 pol intestinalispolyprotein gene, complete cds Genie BK000097 endonuclease TPA_exp:Giardia intestinalis Giardia non-LTR retrotransposon Genie intestinalis1A gene, partial sequence Genie BK000095 endonuclease TPA_exp: Giardiaintestinalis Giardia non-LTR retrotransposon Genie intestinalis 1 gene,partial sequence Genie BK000096 insertion site TPA_exp: Giardiaintestinalis Giardia for non-LTR non-LTR retrotransposon Genieintestinalis retrotransposon 1 target site sequence Genie 1 GenieAY216701 non- Girardia tigrina GENIE Girardia experimentalretrotransposon, complete tigrina evidence, no sequence additionaldetails recorded Genie BK000098 similar to TPA_exp: Giardia intestinalisGiardia endonuclease non-LTR retrotransposon Genie intestinalis 2 gene,complete sequence GilD AF433877 (tca)n (SEQ Giardia intestinalisinactive Giardia ID NO: 1532) non-LTR retrotransposon GilD, intestinalisor (tga)n consensus sequence (SEQ ID NO: 1533), n = 2-4 GilM AF433875poly(dA) Giardia intestinalis non-LTR Giardia tract LINE-likeretrotransposon intestinalis GilM, complete sequence Hero AB097132 rtDanio rerio retrotransposon Danio rerio HERODr DNA, complete sequenceHero AB097130 rt Takifugu rubripes Takifugu rubripes retrotransposonHEROFr DNA, complete sequence HEROTn AB097131 rt Tetraodon nigroviridisTetraodon retrotransposon HEROTn DNA, nigroviridis complete sequenceNeSL_3_135_68117 FJ905846 non-LTR Daphnia pulex non-LTR Daphnia pulexretrotransposon: retrotransposon NeSL_3_135_68117 NeSL_3_135_68117,complete sequence NeSL; DQ099731 target site Caenorhabditis briggsaeCaenorhabditis NeSL-1 duplication transposon NeSl-1-like non-LTRbriggsae retrotransposon NeSL-1Cb reverse transcriptase (pol) gene,complete cds PERERE-9 BN000800 TPA_exp: Schistosoma mansoni SchistosomaPerere-9 non-LTR mansoni retrotransposon R2 AF015814 R2 Limuluspolyphemus Limulus retrotransposon R2, complete polyphemus sequence R2AF090145 R2 Nasonia vitripennis R2 non-LTR Nasonia retrotransposableelement vitripennis reverse transcriptase gene, partial cds R2 AF015818R2 Porcellio scaber Porcellio scaber retrotransposon R2, completesequence R2 AF015815 R2 Anurida maritima Anurida maritimaretrotransposon R2, complete sequence R2 M16558 R2 Bombyx mori rDNAinsertion Bombyx mori element R2 (typeII), complete cds R2 AF015819 R2Forficula auricularia Forficula retrotransposon R2, complete auriculariasequence. R2 EU854578 R2 Triops cancriformis non-LTR Triopsretrotransposon R2 reverse cancriformis transcriptase gene, complete cdsR2 GU949555 R2 Reticulitermes lucifugus Reticulitermes non-LTRretrotransposon R2, lucifugus complete sequence; and R2 protein gene,complete cds R2 AB097123 rt Ciona intestinalis Ciona intestinalisretrotransposon R2Ci-C DNA, partial sequence R2 AB097124 rt Cionaintestinalis Ciona intestinalis retrotransposon R2Ci-D DNA, partialsequence R2 FJ461304 R2 Rhynchosciara americana Rhynchosciara non-LTRretrotransposon RaR2 americana reverse transcriptase gene, complete cdsR2 AB097121 rt Ciona intestinalis Ciona intestinalis retrotransposonR2Ci-A DNA, complete sequence R2 KP657892 R2 Bacillus rossius isolateBacillus rossius roCAP(full).9 retrotransposon R2Br reversetranscriptase gene, partial cds R2 KP657890 R2 Bacillus rossius isolateBacillus rossius roCAP(full).7 retrotransposon R2Br reversetranscriptase gene, partial cds R2 KP657888 R2 Bacillus rossius isolateBacillus rossius roCAP(full).5 retrotransposon R2Br reversetranscriptase gene, partial cds R2 KP657870 R2 Bacillus rossius isolateBacillus rossius roCAP(full).1 retrotransposon R2Br reversetranscriptase gene, partial cds R2 KP657833 R2 Bacillus rossius isolateBacillus rossius roANZ(full).13 retrotransposon R2Br reversetranscriptase gene, partial cds R2 KP657832 R2 Bacillus rossius isolateBacillus rossius roANZ(full).12 retrotransposon R2Br reversetranscriptase gene, partial cds R2 KP657830 R2 Bacillus rossius isolateBacillus rossius roANZ(full).10 retrotransposon R2Br reversetranscriptase gene, partial cds R2 KP657807 R2 Bacillus rossius isolateBacillus rossius roANZ(−101).8 retrotransposon R2Br reversetranscriptase gene, partial cds R2 KP657806 R2 Bacillus rossius isolateBacillus rossius roANZ(−101).7 retrotransposon R2Br reversetranscriptase gene, partial cds R2 KP657805 R2 Bacillus rossius isolateBacillus rossius roANZ(−101).6 retrotransposon R2Br reversetranscriptase gene, partial cds R2 KP657802 R2 Bacillus rossius isolateBacillus rossius roANZ(−101).3 retrotransposon R2Br reversetranscriptase gene, partial cds R2 KP657799 R2 Bacillus rossius isolateBacillus rossius roANZ(−101).1 retrotransposon R2Br reversetranscriptase gene, partial cds R2 FJ461304 R2 Rhynchosciara americanaRhynchosciara non-LTR retrotransposon RaR2 americana reversetranscriptase gene, complete cds R2 JQ082370 polyA_signal_sequenceEyprepocnemis plorans non-LTR Eyprepocnemis retrotransposon R2 R2plorans protein gene, complete cds R2 KJ958672 R2 Bacillus rossiusnon-LTR Bacillus rossius retrotransposon reCUR4_deg, partial sequence R2KJ958671 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonreCUR3_deg, partial sequence R2 KJ958670 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon reCUR2_deg, partial sequence R2KJ958669 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonreCUR1_deg, partial sequence R2 KJ958668 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon reCDF6_deg, partial sequence R2KJ958667 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonreCDF5_deg, partial sequence R2 KJ958666 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon reCDF4_deg, partial sequence R2KJ958665 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonreCDF3_deg, partial sequence R2 KJ958664 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon reCDF2_deg, partial sequence R2KJ958663 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonreCDF1_deg, partial sequence R2 KJ958662 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon reCOM10_deg, partial sequence R2KJ958661 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonreCOM5_deg, partial sequence R2 KJ958660 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon reCOM7_deg, partial sequence R2KJ958659 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonreCOM3_deg, partial sequence R2 KJ958658 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon reCOM2_deg, partial sequence R2KJ958657 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonreMSN6_deg, partial sequence R2 KJ958656 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon reMSN5_deg, partial sequence R2KJ958655 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonreMSN4_deg, partial sequence R2 KJ958654 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon reMSN3_deg, partial sequence R2KJ958653 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonreMSN2_deg, partial sequence R2 KJ958652 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon reMSN1_deg, partial sequence R2KJ958651 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonrePAT8_deg, partial sequence R2 KJ958650 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon rePAT7_deg, partial sequence R2KJ958649 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonrePAT6_deg, partial sequence R2 KJ958648 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon rePAT5_deg, partial sequence R2KJ958647 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonrePAT4_deg, partial sequence R2 KJ958646 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon rePAT3_deg, partial sequence R2KJ958645 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonrePAT2_deg, partial sequence R2 KJ958644 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon rePAT1_deg, partial sequence R2KJ958643 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonrePAT9_deg, partial sequence R2 KJ958642 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon trKOR2_deg, partial sequence R2KJ958641 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonreGAB9_deg, partial sequence R2 KJ958640 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon reGAB8_deg, partial sequence R2KJ958639 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonreGAB7_deg, partial sequence R2 KJ958638 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon reGAB6_deg, partial sequence R2KJ958637 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonreGAB5_deg, partial sequence R2 KJ958636 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon reGAB4_deg, partial sequence R2KJ958635 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonreGAB3_deg, partial sequence R2 KJ958634 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon reGAB2_deg, partial sequence R2KJ958633 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonreGAB1_deg, partial sequence R2 KJ958632 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon reTDS1_deg, partial sequence R2KJ958631 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonreTDS11_deg, partial sequence R2 KJ958629 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon reCOM8, partial sequence R2 KJ958628 R2Bacillus rossius non-LTR Bacillus rossius retrotransposon reCOM6,partial sequence R2 KJ958627 R2 Bacillus rossius non-LTR Bacillusrossius retrotransposon reCOM4, partial sequence R2 KJ958626 R2 Bacillusrossius non-LTR Bacillus rossius retrotransposon reCOM1, partialsequence R2 KJ958624 R2 Bacillus rossius non-LTR Bacillus rossiusretrotransposon trKOR10, partial sequence R2 KJ958623 R2 Bacillusrossius non-LTR Bacillus rossius retrotransposon reGAB10, partialsequence R2 KJ958619 R2 Bacillus rossius non-LTR Bacillus rossiusretrotransposon trKOR4, partial sequence R2 KJ958618 R2 Bacillus rossiusnon-LTR Bacillus rossius retrotransposon trKOR3, partial sequence R2KJ958617 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposontrKOR1, partial sequence R2 KJ958613 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon reVIR4, partial sequence R2 KJ958612 R2Bacillus rossius non-LTR Bacillus rossius retrotransposon reVIR3,partial sequence R2 KJ958611 R2 Bacillus rossius non-LTR Bacillusrossius retrotransposon reVIR2, partial sequence R2 KJ958610 R2 Bacillusrossius non-LTR Bacillus rossius retrotransposon reVIR1, partialsequence R2 KJ958609 R2 Bacillus rossius non-LTR Bacillus rossiusretrotransposon reTDS17, partial sequence R2 KJ958608 R2 Bacillusrossius non-LTR Bacillus rossius retrotransposon reTDS16, partialsequence R2 KJ958607 R2 Bacillus rossius non-LTR Bacillus rossiusretrotransposon reTDS15, partial sequence R2 KJ958606 R2 Bacillusrossius non-LTR Bacillus rossius retrotransposon reTDS14, partialsequence R2 KJ958605 R2 Bacillus rossius non-LTR Bacillus rossiusretrotransposon reTDS13, partial sequence R2 KJ958604 R2 Bacillusrossius non-LTR Bacillus rossius retrotransposon reTDS12, partialsequence R2 KJ958603 R2 Bacillus rossius non-LTR Bacillus rossiusretrotransposon reTDS7, partial sequence R2 KJ958602 R2 Bacillus rossiusnon-LTR Bacillus rossius retrotransposon reTDS6, partial sequence R2KJ958601 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonreTDS5, partial sequence R2 KJ958600 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon reTDS4, partial sequence R2 KJ958599 R2Bacillus rossius non-LTR Bacillus rossius retrotransposon reTDS3,partial sequence R2 KJ958598 R2 Bacillus rossius non-LTR Bacillusrossius retrotransposon reTDS2, partial sequence R2 KJ958594 R2 Bacillusrossius non-LTR Bacillus rossius retrotransposon reBER4, partialsequence R2 KJ958593 R2 Bacillus rossius non-LTR Bacillus rossiusretrotransposon reBER2, partial sequence R2 KJ958592 R2 Bacillus rossiusnon-LTR Bacillus rossius retrotransposon reBER1, partial sequence R2KJ958591 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonroFOL7, partial sequence R2 KJ958590 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon roFOL6, partial sequence R2 KJ958589 R2Bacillus rossius non-LTR Bacillus rossius retrotransposon roFOL5,partial sequence R2 KJ958588 R2 Bacillus rossius non-LTR Bacillusrossius retrotransposon roFOL4, partial sequence R2 KJ958587 R2 Bacillusrossius non-LTR Bacillus rossius retrotransposon roFOL3, partialsequence R2 KJ958586 R2 Bacillus rossius non-LTR Bacillus rossiusretrotransposon roFOL2, partial sequence R2 KJ958585 R2 Bacillus rossiusnon-LTR Bacillus rossius retrotransposon roFOL1, partial sequence R2KJ958584 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonroANZ10, partial sequence R2 KJ958583 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon roANZ9, partial sequence R2 KJ958582 R2Bacillus rossius non-LTR Bacillus rossius retrotransposon roANZ8,partial sequence R2 KJ958581 R2 Bacillus rossius non-LTR Bacillusrossius retrotransposon roANZ7, partial sequence R2 KJ958580 R2 Bacillusrossius non-LTR Bacillus rossius retrotransposon roANZ6, partialsequence R2 KJ958579 R2 Bacillus rossius non-LTR Bacillus rossiusretrotransposon roANZ5, partial sequence R2 KJ958578 R2 Bacillus rossiusnon-LTR Bacillus rossius retrotransposon roANZ4, partial sequence R2KJ958577 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonroANZ3, partial sequence R2 KJ958576 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon roANZ2, partial sequence R2 KJ958575 R2Bacillus rossius non-LTR Bacillus rossius retrotransposon roANZ1,partial sequence R2 KJ958574 R2 Bacillus rossius non-LTR Bacillusrossius retrotransposon roCAP9, partial sequence R2 KJ958573 R2 Bacillusrossius non-LTR Bacillus rossius retrotransposon roCAP8, partialsequence R2 KJ958572 R2 Bacillus rossius non-LTR Bacillus rossiusretrotransposon roCAP7, partial sequence R2 KJ958571 R2 Bacillus rossiusnon-LTR Bacillus rossius retrotransposon roCAP6, partial sequence R2KJ958570 R2 Bacillus rossius non-LTR Bacillus rossius retrotransposonroCAP5, partial sequence R2 KJ958569 R2 Bacillus rossius non-LTRBacillus rossius retrotransposon roCAP4, partial sequence R2 KJ958568 R2Bacillus rossius non-LTR Bacillus rossius retrotransposon roCAP3,partial sequence R2 KJ958567 R2 Bacillus rossius non-LTR Bacillusrossius retrotransposon roCAP2, partial sequence R2 KJ958566 R2 Bacillusrossius non-LTR Bacillus rossius retrotransposon roCAP1, partialsequence R2 KJ958565 R2 Bacillus rossius non-LTR Bacillus rossiusretrotransposon roCAP10, partial sequence R2 JN937654 R2 Lepidurus apuslubbocki Lepidurus apus isolate lu8a 28S ribosomal lubbocki RNA gene,partial sequence; and non-LTR retrotransposon psR2Ll, complete sequenceR2 JN937653 R2 Lepidurus apus lubbocki Lepidurus apus isolate lu7a 28Sribosomal lubbocki RNA gene, partial sequence; and non-LTRretrotransposon psR2Ll, complete sequence R2 JN937652 R2 Lepidurus apuslubbocki Lepidurus apus isolate lu2a 28S ribosomal lubbocki RNA gene,partial sequence; and non-LTR retrotransposon psR2Ll, complete sequenceR2 JN937651 R2 Lepidurus apus lubbocki Lepidurus apus isolate b7c7 28Sribosomal lubbocki RNA gene, partial sequence; and non-LTRretrotransposon psR2Ll, complete sequence R2 JN937650 R2 Lepidurus apuslubbocki Lepidurus apus isolate b6c4 28S ribosomal lubbocki RNA gene,partial sequence; and non-LTR retrotransposon psR2Ll, complete sequenceR2 JN937649 R2 Lepidurus apus lubbocki Lepidurus apus isolate lu5a 28Sribosomal lubbocki RNA gene, partial sequence; and non-LTRretrotransposon psR2Ll, complete sequence R2 JN937648 R2 Lepidurus apuslubbocki Lepidurus apus isolate lu1a 28S ribosomal lubbocki RNA gene,partial sequence; and non-LTR retrotransposon psR2Ll, complete sequenceR2 JN937647 R2 Lepidurus apus lubbocki Lepidurus apus isolate b6c5 28Sribosomal lubbocki RNA gene, partial sequence; and non-LTRretrotransposon psR2Ll, complete sequence R2 JN937646 R2 Lepidurus apuslubbocki Lepidurus apus isolate b6c6 28S ribosomal lubbocki RNA gene,partial sequence; and non-LTR retrotransposon psR2Ll, complete sequenceR2 JN937645 R2 Lepidurus apus lubbocki Lepidurus apus isolate lu4a 28Sribosomal lubbocki RNA gene, partial sequence; and non-LTRretrotransposon psR2Ll, complete sequence R2 JN937644 R2 Lepidurus apuslubbocki Lepidurus apus isolate lu3a 28S ribosomal lubbocki RNA gene,partial sequence; and non-LTR retrotransposon psR2Ll, complete sequenceR2 JN937643 R2 Lepidurus apus lubbocki Lepidurus apus isolate b6c3 28Sribosomal lubbocki RNA gene, partial sequence; and non-LTRretrotransposon psR2Ll, complete sequence R2 JN937642 R2 Lepidurus apuslubbocki Lepidurus apus isolate lu6a 28S ribosomal lubbocki RNA gene,partial sequence; and non-LTR retrotransposon psR2Ll, complete sequenceR2 JN937641 R2 Lepidurus apus lubbocki Lepidurus apus isolate LM5h2 28Sribosomal lubbocki RNA gene, partial sequence; and non-LTRretrotransposon R2Ll, complete sequence R2 JN937640 R2 Lepidurus apuslubbocki Lepidurus apus isolate LM2h5 28S ribosomal lubbocki RNA gene,partial sequence; and non-LTR retrotransposon R2Ll, complete sequence R2JN937639 R2 Lepidurus apus lubbocki Lepidurus apus isolate LM2h4 28Sribosomal lubbocki RNA gene, partial sequence; and non-LTRretrotransposon R2Ll, complete sequence R2 JN937638 R2 Lepidurus apuslubbocki Lepidurus apus isolate LM5h5 28S ribosomal lubbocki RNA gene,partial sequence; and non-LTR retrotransposon R2Ll, complete sequence R2JN937637 R2 Lepidurus apus lubbocki Lepidurus apus isolate LM5h4 28Sribosomal lubbocki RNA gene, partial sequence; and non-LTRretrotransposon R2Ll, complete sequence R2 JN937636 R2 Lepidurus apuslubbocki Lepidurus apus isolate LM5h3 28S ribosomal lubbocki RNA gene,partial sequence; and non-LTR retrotransposon R2Ll, complete sequence R2JN937635 R2 Lepidurus apus lubbocki Lepidurus apus isolate LM5h1 28Sribosomal lubbocki RNA gene, partial sequence; and non-LTRretrotransposon R2Ll, complete sequence R2 JN937634 R2 Lepidurus apuslubbocki Lepidurus apus isolate LM2h3 28S ribosomal lubbocki RNA gene,partial sequence; and non-LTR retrotransposon R2Ll, complete sequence R2JN937633 R2 Lepidurus apus lubbocki Lepidurus apus isolate LM2h2 28Sribosomal lubbocki RNA gene, partial sequence; and non-LTRretrotransposon R2Ll, complete sequence R2 JN937632 R2 Lepidurus apuslubbocki Lepidurus apus isolate LM2h1 28S ribosomal lubbocki RNA gene,partial sequence; and non-LTR retrotransposon R2Ll, complete sequence R2JN937631 R2 Lepidurus arcticus isolate Lepidurus arcticus T6 28Sribosomal RNA gene, partial sequence; and non-LTR retrotransposon R2La,complete sequence R2 JN937630 R2 Lepidurus arcticus isolate Lepidurusarcticus T5 28S ribosomal RNA gene, partial sequence; and non-LTRretrotransposon R2La, complete sequence R2 JN937629 R2 Lepidurusarcticus isolate Lepidurus arcticus T4 28S ribosomal RNA gene, partialsequence; and non-LTR retrotransposon R2La, complete sequence R2JN937628 R2 Lepidurus arcticus isolate Lepidurus arcticus T3 28Sribosomal RNA gene, partial sequence; and non-LTR retrotransposon R2La,complete sequence R2 JN937627 R2 Lepidurus arcticus isolate Lepidurusarcticus T2 28S ribosomal RNA gene, partial sequence; and non-LTRretrotransposon R2La, complete sequence R2 JN937626 R2 Lepidurusarcticus isolate Lepidurus arcticus T1 28S ribosomal RNA gene, partialsequence; and non-LTR retrotransposon R2La, complete sequence R2JN937625 R2 Lepidurus arcticus isolate Lepidurus arcticus V4 28Sribosomal RNA gene, partial sequence; and non-LTR retrotransposon R2La,complete sequence R2 JN937624 R2 Lepidurus arcticus isolate Lepidurusarcticus V3 28S ribosomal RNA gene, partial sequence; and non-LTRretrotransposon R2La, complete sequence R2 JN937623 R2 Lepidurusarcticus isolate Lepidurus arcticus V2 28S ribosomal RNA gene, partialsequence; and non-LTR retrotransposon R2La, complete sequence R2JN937622 R2 Lepidurus arcticus isolate Lepidurus arcticus V1 28Sribosomal RNA gene, partial sequence; and non-LTR retrotransposon R2La,complete sequence R2 JN937615 R2 Lepidurus couesii isolate Lepiduruscouesii D3a7f 28S ribosomal RNA gene, partial sequence; and non-LTRretrotransposon R2LcB, complete sequence R2 JN937614 R2 Lepiduruscouesii isolate Lepidurus couesii D3a5f 28S ribosomal RNA gene, partialsequence; and non-LTR retrotransposon R2LcB, complete sequence R2JN937613 R2 Lepidurus couesii isolate Lepidurus couesii D3a4f 28Sribosomal RNA gene, partial sequence; and non-LTR retrotransposon R2LcB,complete sequence R2 JN937612 R2 Lepidurus couesii isolate Lepiduruscouesii D3a3f 28S ribosomal RNA gene, partial sequence; and non-LTRretrotransposon R2LcB, complete sequence R2 JN937611 R2 Lepiduruscouesii isolate Lepidurus couesii D3a2f 28S ribosomal RNA gene, partialsequence; and non-LTR retrotransposon R2LcB, complete sequence R2JN937610 R2 Lepidurus couesii isolate Lepidurus couesii D3_8 28Sribosomal RNA gene, partial sequence; and non-LTR retrotransposon R2LcB,complete sequence R2 JN937609 R2 Lepidurus couesii isolate Lepiduruscouesii D3_7 28S ribosomal RNA gene, partial sequence; and non-LTRretrotransposon R2LcB, complete sequence R2 JN937608 R2 Lepiduruscouesii isolate Lepidurus couesii D3_6 28S ribosomal RNA gene, partialsequence; and non-LTR retrotransposon R2LcB, complete sequence R2JN937607 R2 Lepidurus couesii isolate Lepidurus couesii D3_5 28Sribosomal RNA gene, partial sequence; and non-LTR retrotransposon R2LcB,complete sequence R2 JN937606 R2 Lepidurus couesii isolate Lepiduruscouesii D3_4 28S ribosomal RNA gene, partial sequence; and non-LTRretrotransposon R2LcB, complete sequence R2 JN937605 R2 Lepiduruscouesii isolate Lepidurus couesii D3_3 28S ribosomal RNA gene, partialsequence; and non-LTR retrotransposon R2LcB, complete sequence R2JN937604 R2 Lepidurus couesii isolate Lepidurus couesii D3_2 28Sribosomal RNA gene, partial sequence; and non-LTR retrotransposon R2LcB,complete sequence R2 JN937603 R2 Lepidurus couesii isolate Lepiduruscouesii D3_1 28S ribosomal RNA gene, partial sequence; and non-LTRretrotransposon R2LcB, complete sequence R2 JN937602 R2 Lepiduruscouesii isolate Lepidurus couesii C2_5 28S ribosomal RNA gene, partialsequence; and non-LTR retrotransposon R2LcA, complete sequence R2JN937601 R2 Lepidurus couesii isolate Lepidurus couesii LcoC2r1_5 28Sribosomal RNA gene, partial sequence; and non-LTR retrotransposon R2LcA,complete sequence R2 JN937600 R2 Lepidurus couesii isolate Lepiduruscouesii LcoC2r1_6 28S ribosomal RNA gene, partial sequence; and non-LTRretrotransposon R2LcA, complete sequence R2 JN937599 R2 Lepiduruscouesii isolate Lepidurus couesii C2_8 28S ribosomal RNA gene, partialsequence; and non-LTR retrotransposon R2LcA, complete sequence R2JN937598 R2 Lepidurus couesii isolate Lepidurus couesii C2_4 28Sribosomal RNA gene, partial sequence; and non-LTR retrotransposon R2LcA,complete sequence R2 JN937597 R2 Lepidurus couesii isolate Lepiduruscouesii C2_9 28S ribosomal RNA gene, partial sequence; and non-LTRretrotransposon R2LcA, complete sequence R2 JN937596 R2 Lepiduruscouesii isolate Lepidurus couesii C2_7 28S ribosomal RNA gene, partialsequence; and non-LTR retrotransposon R2LcA, complete sequence R2JN937595 R2 Lepidurus couesii isolate Lepidurus couesii C2_6 28Sribosomal RNA gene, partial sequence; and non-LTR retrotransposon R2LcA,complete sequence R2 JN937594 R2 Lepidurus couesii isolate Lepiduruscouesii C2_3 28S ribosomal RNA gene, partial sequence; and non-LTRretrotransposon R2LcA, complete sequence R2 JN937593 R2 Lepiduruscouesii isolate Lepidurus couesii C2_2 28S ribosomal RNA gene, partialsequence; and non-LTR retrotransposon R2LcA, complete sequence R2JN937592 R2 Lepidurus couesii isolate Lepidurus couesii C2_1 28Sribosomal RNA gene, partial sequence; and non-LTR retrotransposon R2LcA,complete sequence R2 AF015822 R2 ORF Tenebrio molitor Tenebrio molitorretrotransposon R2 reverse transcriptase gene, partial cds R2 AF015817R2 ORF Tenebrio molitor Tenebrio molitor retrotransposon R2 reversetranscriptase gene, partial cds R2 AF015816 R2 ORF Hippodamia convergensHippodamia retrotransposon R2 reverse convergens transcriptase gene,partial cds R2 KP657866 retrotransposon: Bacillus rossius isolateBacillus rossius R2Br roCAP(−714).5 retrotransposon R2Br reversetranscriptase gene, partial cds R2 KP657865 retrotransposon: Bacillusrossius isolate Bacillus rossius R2Br roCAP(−714).4 retrotransposon R2Brreverse transcriptase gene, partial cds R2 KP657863 retrotransposon:Bacillus rossius isolate Bacillus rossius R2Br roCAP(−714).2retrotransposon R2Br reverse transcriptase gene, partial cds R2 KP657862retrotransposon: Bacillus rossius isolate Bacillus rossius R2BrroCAP(−714).1 retrotransposon R2Br reverse transcriptase gene, partialcds R2 KP657861 retrotransposon: Bacillus rossius isolate Bacillusrossius R2Br roCAP(−1297).9 retrotransposon R2Br reverse transcriptasegene, partial cds R2 KP657860 retrotransposon: Bacillus rossius isolateBacillus rossius R2Br roCAP(−1297).8 retrotransposon R2Br reversetranscriptase gene, partial cds R2 KP657859 retrotransposon: Bacillusrossius isolate Bacillus rossius R2Br roCAP(−1297).7 retrotransposonR2Br reverse transcriptase gene, partial cds R2 KP657858retrotransposon: Bacillus rossius isolate Bacillus rossius R2BrroCAP(−1297).6 retrotransposon R2Br reverse transcriptase gene, partialcds R2 KP657857 retrotransposon: Bacillus rossius isolate Bacillusrossius R2Br roCAP(−1297).5 retrotransposon R2Br reverse transcriptasegene, partial cds R2 KP657856 retrotransposon: Bacillus rossius isolateBacillus rossius R2Br roCAP(−1297).4 retrotransposon R2Br reversetranscriptase gene, partial cds R2 KP657855 retrotransposon: Bacillusrossius isolate Bacillus rossius R2Br roCAP(−1297).3 retrotransposonR2Br reverse transcriptase gene, partial cds R2 KP657854retrotransposon: Bacillus rossius isolate Bacillus rossius R2BrroCAP(−1297).2 retrotransposon R2Br reverse transcriptase gene, partialcds R2 KP657853 retrotransposon: Bacillus rossius isolate Bacillusrossius R2Br roCAP(−1297).10 retrotransposon R2Br reverse transcriptasegene, partial cds R2 KP657852 retrotransposon: Bacillus rossius isolateBacillus rossius R2Br roCAP(−1297).1 retrotransposon R2Br reversetranscriptase gene, partial cds R2 KP657851 retrotransposon: Bacillusrossius isolate Bacillus rossius R2Br roCAP(−1172).9 retrotransposonR2Br reverse transcriptase gene, partial cds R2 KP657850retrotransposon: Bacillus rossius isolate Bacillus rossius R2BrroCAP(−1172).8 retrotransposon R2Br reverse transcriptase gene, partialcds R2 KP657849 retrotransposon: Bacillus rossius isolate Bacillusrossius R2Br roCAP(−1172).7 retrotransposon R2Br reverse transcriptasegene, partial cds R2 KP657848 retrotransposon: Bacillus rossius isolateBacillus rossius R2Br roCAP(−1172).6 retrotransposon R2Br reversetranscriptase gene, partial cds R2 KP657847 retrotransposon: Bacillusrossius isolate Bacillus rossius R2Br roCAP(−1172).5 retrotransposonR2Br reverse transcriptase gene, partial cds R2 KP657846retrotransposon: Bacillus rossius isolate Bacillus rossius R2BrroCAP(−1172).4 retrotransposon R2Br reverse transcriptase gene, partialcds R2 KP657845 retrotransposon: Bacillus rossius isolate Bacillusrossius R2Br roCAP(−1172).3 retrotransposon R2Br reverse transcriptasegene, partial cds R2 KP657844 retrotransposon: Bacillus rossius isolateBacillus rossius R2Br roCAP(−1172).2 retrotransposon R2Br reversetranscriptase gene, partial cds R2 KP657843 retrotransposon: Bacillusrossius isolate Bacillus rossius R2Br roCAP(−1172).10 retrotransposonR2Br reverse transcriptase gene, partial cds R2 KP657842retrotransposon: Bacillus rossius isolate Bacillus rossius R2BrroCAP(−1172).1 retrotransposon R2Br reverse transcriptase gene, partialcds R2 KP657824 retrotransposon: Bacillus rossius isolate Bacillusrossius R2Br roANZ(−1062).5 retrotransposon R2Br reverse transcriptasegene, partial cds R2 KP657823 retrotransposon: Bacillus rossius isolateBacillus rossius R2Br roANZ(−1062).4 retrotransposon R2Br reversetranscriptase gene, partial cds R2 KP657822 retrotransposon: Bacillusrossius isolate Bacillus rossius R2Br roANZ(−1062).3 retrotransposonR2Br reverse transcriptase gene, partial cds R2 KP657820retrotransposon: Bacillus rossius isolate Bacillus rossius R2BrroANZ(−1062).2 retrotransposon R2Br reverse transcriptase gene, partialcds R2 KP657816 retrotransposon: Bacillus rossius isolate Bacillusrossius R2Br roANZ(−1062).16 retrotransposon R2Br reverse transcriptasegene, partial cds R2 KP657814 retrotransposon: Bacillus rossius isolateBacillus rossius R2Br roANZ(−1062).14 retrotransposon R2Br reversetranscriptase gene, partial cds R2 KP657810 retrotransposon: Bacillusrossius isolate Bacillus rossius R2Br roANZ(−1062).10 retrotransposonR2Br reverse transcriptase gene, partial cds R2 KP657809retrotransposon: Bacillus rossius isolate Bacillus rossius R2BrroANZ(−1062).1 retrotransposon R2Br reverse transcriptase gene, partialcds R2 KP657759 retrotransposon: Bacillus rossius isolate Bacillusrossius R2Br rePAT(−1297).8 retrotransposon R2Br reverse transcriptasegene, partial cds R2 KP657757 retrotransposon: Bacillus rossius isolateBacillus rossius R2Br rePAT(−1297).6 retrotransposon R2Br reversetranscriptase gene, partial cds R2 KP657751 retrotransposon: Bacillusrossius isolate Bacillus rossius R2Br rePAT(−1297).1 retrotransposonR2Br reverse transcriptase gene, partial cds R2 AB097125 rt Cionasavignyi Ciona savignyi retrotransposon R2Cs-D DNA, partial sequence R2AB097124 rt Ciona intestinalis Ciona intestinalis retrotransposon R2Ci-DDNA, partial sequence R2 AB097123 rt Ciona intestinalis Cionaintestinalis retrotransposon R2Ci-C DNA, partial sequence R2 AB097121 rtCiona intestinalis Ciona intestinalis retrotransposon R2Ci-A DNA,complete sequence R2 AB201417 rt Triops longicaudatus non-LTR Triopsretrotransposon R2Tl gene longicaudatus for reverse transcriptase,partial cds R2 AB201416 rt Procambarus clarkii non-LTR Procambarusretrotransposon R2Pc gene for clarkii reverse transcriptase, partial cdsR2 AB201415 rt Hasarius adansoni non-LTR Hasarius adansoniretrotransposon R2Ha gene for reverse transcriptase, partial cds R2AB201414 rt Metacrinus rotundus non-LTR Metacrinus retrotransposon R2Mrgene for rotundus reverse transcriptase, partial cds R2 AB201413 rtMauremys reevesii non-LTR Mauremys reevesii retrotransposon R2Cr-B2 genefor reverse transcriptase, partial cds R2 AB201412 rt Mauremys reevesiinon-LTR Mauremys reevesii retrotransposon R2Cr-B1 gene for reversetranscriptase, partial cds R2 AB201411 rt Mauremys reevesii non-LTRMauremys reevesii retrotransposon R2Cr-A gene for reverse transcriptase,partial cds R2 AB201410 rt Oryzias latipes non-LTR Oryzias latipesretrotransposon R2Ol-A gene for reverse transcriptase, partial cds R2AB201409 rt Tanichthys albonubes non-LTR Tanichthys retrotransposon R2Tagene albonubes for reverse transcriptase, partial cds R2 AB201408 rtEptatretus burgeri non-LTR Eptatretus retrotransposon R2Eb gene forburgeri reverse transcriptase, partial cds R2 DQ099732 transposon: Aedesaegypti transposon R2- Aedes aegypti R2-like like non-LTRretrotransposon non-LTR R2Ag reverse transcriptase retrotransposon (pol)gene, partial cds R2Ag R2 DQ099728 transposon: Aedes aegypti transposonR2- Aedes aegypti R2-like like non-LTR retrotransposon non-LTR R2Ag_Breverse transcriptase retrotransposon (pol) gene, partial cds R2Ag_B R2GU949559 Kalotermes flavicollis Kalotermes isolate Livorno non-LTRflavicollis retrotransposon R2, complete sequence; and R2 protein gene,complete cds R2 GU949557 Reticulitermes balkanensis Reticulitermesnon-LTR retrotransposon R2, balkanensis partial sequence; and R2 proteingene, partial cds R2 GU949556 Reticulitermes grassei Reticulitermesnon-LTR retrotransposon R2, grassei partial sequence; and R2 proteingene, partial cds R2 GU949554 Reticulitermes urbis non-LTRReticulitermes retrotransposon R2, complete urbis sequence; and R2protein gene, complete cds R2 AF412214 Schistosoma japonicum cloneSchistosoma S10A non-LTR retrotransposon japonicum SjR2-like, partialsequence R2 AF015685 Drosophila mercatorum R2 Drosophila retrotransposonreverse mercatorum transcriptase domain protein gene, complete cds R2KJ958674 Bacillus rossius Bacillus rossius retrotransposon R2Br,complete sequence R2 AF015814 R2 Limulus polyphemus Limulus polyphemusretrotransposon R2, complete sequence R2 M16558 R2 Bombyx mori rDNAinsertion Bombyx mori element R2 (typeII), complete cds. R2 GQ398057R9Av Adineta vaga copy 1 non-LTR Adineta vaga retrotransposon R9,complete sequence; and disrupted 28S ribosomal RNA gene, partialsequence R2Bm AB076841 R2 Bombyx mori non-LTR Bombyx moriretrotransposon R2Bm gene for reverse transcriptase, complete cds and28S rRNA R2Ci-B AB097122 rt Ciona intestinalis Ciona intestinalisretrotransposon R2Ci-B DNA, complete sequence R2Dr AB097126 rt Daniorerio retrotransposon Danio rerio R2Dr DNA, complete sequence R4AH003588 Parascaris equorum transposon Parascaris equorum non-LTRretrotransposable element R4 reverse transcriptase gene, partial cds R4ALU29445 R4 Ascaris lumbricoides Ascaris site-specific non-LTRlumbricoides retrotransposable element R4 in 26S rDNA, complete sequenceR4 L08889 R4 Dong Bombyx mori reverse Bombyx mori transcriptase gene,complete cds R4 DQ836390 MalR4-5 Maculinea alcon R4-like Phengaris alconnon-LTR retrotransposon R4-5 reverse transcriptase (RT) pseudogene,partial sequence R4 DQ836385 MnaR4-3 Maculinea nausithous R4-likePhengaris non-LTR retrotransposon R4-3 nausithous reverse transcriptase(RT) pseudogene, partial sequence R4 DQ836386 MnaR4-4 Maculineanausithous R4-like Phengaris non-LTR retrotransposon R4-4 nausithousreverse transcriptase (RT) pseudogene, partial sequence R4 DQ836387MnaR4-7 Maculinea nausithous R4-like Phengaris non-LTR retrotransposonR4-7 nausithous reverse transcriptase (RT) pseudogene, partial sequenceR4 DQ836388 MnaR4-8 Maculinea nausithous R4-like Phengaris non-LTRretrotransposon R4-8 nausithous reverse transcriptase (RT) pseudogene,partial sequence R4 DQ836389 MnaR4-9 Maculinea nausithous R4-likePhengaris non-LTR retrotransposon R4-9 nausithous reverse transcriptase(RT) pseudogene, partial sequence R4 DQ836379 MteR4-1 Maculinea teleiusR4-like Phengaris teleius non-LTR retrotransposon R4-1 reversetranscriptase (RT) pseudogene, partial sequence R4 DQ836384 MteR4-10Maculinea teleius R4-like Phengaris teleius non-LTR retrotransposonR4-10 reverse transcriptase (RT) pseudogene, partial sequence R4DQ836367 MteR4-2 Maculinea teleius R4-like Phengaris teleius non-LTRretrotransposon R4-2 reverse transcriptase (RT) gene, partial cds R4DQ836380 MteR4-3 Maculinea teleius R4-like Phengaris teleius non-LTRretrotransposon R4-3 reverse transcriptase (RT) pseudogene, partialsequence R4 DQ836381 MteR4-4 Maculinea teleius R4-like Phengaris teleiusnon-LTR retrotransposon R4-4 reverse transcriptase (RT) pseudogene,partial sequence R4 DQ836382 MteR4-6 Maculinea teleius R4-like Phengaristeleius non-LTR retrotransposon R4-6 reverse transcriptase (RT)pseudogene, partial sequence R4 DQ836383 MteR4-8 Maculinea teleiusR4-like Phengaris teleius non-LTR retrotransposon R4-8 reversetranscriptase (RT) pseudogene, partial sequence R4 DQ836374 transposon:Maculinea alcon R4-like Phengaris alcon R4-like non-LTR retrotransposonR4-1 non-LTR reverse transcriptase (RT) retrotransposon gene, partialcds R4-1 R4 DQ836373 transposon: Maculinea nausithous R4-like PhengarisR4-like non-LTR retrotransposon R4-1 nausithous non-LTR reversetranscriptase (RT) retrotransposon gene, partial cds R4-1 R4 DQ836375transposon: Maculinea alcon R4-like Phengaris alcon R4-like non-LTRretrotransposon R4-2 non-LTR reverse transcriptase (RT) retrotransposongene, partial cds R4-2 R4 DQ836376 transposon: Maculinea alcon R4-likePhengaris alcon R4-like non-LTR retrotransposon R4-3 non-LTR reversetranscriptase (RT) retrotransposon gene, partial cds R4-3 R4 DQ836377transposon: Maculinea alcon R4-like Phengaris alcon R4-like non-LTRretrotransposon R4-4 non-LTR reverse transcriptase (RT) retrotransposongene, partial cds R4-4 R4 DQ836371 transposon: Maculinea nausithousR4-like Phengaris R4-like non-LTR retrotransposon R4-5 nausithousnon-LTR reverse transcriptase (RT) retrotransposon gene, partial cdsR4-5 R4 DQ836368 transposon: Maculinea teleius R4-like Phengaris teleiusR4-like non-LTR retrotransposon R4-5 non-LTR reverse transcriptase (RT)retrotransposon gene, partial cds R4-5 R4 DQ836378 transposon: Maculineaalcon R4-like Phengaris alcon R4-like non-LTR retrotransposon R4-6non-LTR reverse transcriptase (RT) retrotransposon gene, partial cdsR4-6 R4 DQ836372 transposon: Maculinea nausithous R4-like PhengarisR4-like non-LTR retrotransposon R4-6 nausithous non-LTR reversetranscriptase (RT) retrotransposon gene, partial cds R4-6 R4 DQ836369transposon: Maculinea teleius R4-like Phengaris teleius R4-like non-LTRretrotransposon R4-7 non-LTR reverse transcriptase (RT) retrotransposongene, partial cds R4-7 R4 DQ836370 transposon: Maculinea teleius R4-likePhengaris teleius R4-like non-LTR retrotransposon R4-9 non-LTR reversetranscriptase (RT) retrotransposon gene, partial cds R4-9 R4 AF286191transposon: Xiphophorus maculatus Xiphophorus retrotransposonretrotransposon Rex6 reverse maculatus Rex6 transcriptase pseudogene,partial sequence R5 KJ958673 R2 Bacillus rossius non-LTR Bacillusrossius retrotransposon reCUR5_deg, partial sequence R5 KJ958620 R2Bacillus rossius non-LTR Bacillus rossius retrotransposon trKOR5,partial sequence R5 KJ958614 R2 Bacillus rossius non-LTR Bacillusrossius retrotransposon reVIR5, partial sequence R5 KJ958595 R2 Bacillusrossius non-LTR Bacillus rossius retrotransposon reBER5, partialsequence R5 AJ006560 transposon: Anopheles merus Amer5 non-LTR Anophelesmerus Amer5 retrotransposon encoding non-LTR reverse transcriptase,partial retrotransposon R5 AF352479 transposon: Chironomus circumdatusclone Chironomus NLRCth1-like cir5 transposon NLRCth1-like circumdatusnon-LTR non-LTR retrotransposon retrotransposon reverse transcriptasegene, partial cds R5 AF352454 transposon: Chironomus alpestris cloneChironomus NLRCth1-like dor50 transposon NLRCth1-like alpestris non-LTRnon-LTR retrotransposon retrotransposon reverse transcriptase gene,partial cds R5 AF352404 transposon: Chironomus luridus clone lur5Chironomus luridus NLRCth1-like transposon NLRCth1-like non-LTR non-LTRretrotransposon retrotransposon reverse transcriptase gene, partial cdsR8 FR852798 poly(A) tail Beta vulgaris subsp. Beta vulgaris vulgarissubsp. vulgaris LINE-type retrotransposon Belline2_3 R8 KJ958630 R2Bacillus rossius non-LTR Bacillus rossius retrotransposon reVIR8_deg,partial sequence R8 KJ958621 R2 Bacillus rossius non-LTR Bacillusrossius retrotransposon trKOR8, partial sequence R8 KP001560 rex3-RTIberochondrostoma lusitanicum Iberochondrostoma pseudogene_Contig clonetr8a non-LTR lusitanicum ILU_TR8 retrotransposon Rex3, complete sequenceR8 FR852885 right terminal Beta vulgaris subsp. Beta vulgaris repeatvulgaris subsp. vulgaris LINE-type retrotransposon BNR114 (Belline1_114)R8 FR852856 right terminal Beta vulgaris subsp. Beta vulgaris repeatvulgarius subsp. vulgaris LINE-type retrotransposon BNR45 (Belline1_45)R8 FR852844 right terminal Beta vulgaris subsp. Beta vulgaris repeatvulgarius subsp. vulgaris LINE-type retrotransposon BNR22 (Belline1_22)R8 FR852836 right terminal Beta vulgaris subsp. Beta vulgaris repeatvulgarius subsp. vulgaris LINE-type retrotransposon Belline17_6 R8FR852834 right terminal Beta vulgaris subsp. Beta vulgaris repeatvulgarius subsp. vulgaris LINE-type retrotransposon Belline17_4 R8FR852831 right terminal Beta vulgaris subsp. Beta vulgaris repeatvulgarius subsp. vulgaris LINE-type retrotransposon Belline17_1 R8FR852829 right terminal Beta vulgaris subsp. Beta vulgaris repeatvulgarius subsp. vulgaris LINE-type retrotransposon Belline16_2 R8FR852827 right terminal Beta vulgaris subsp. Beta vulgaris repeatvulgarius subsp. vulgaris LINE-type retrotransposon Belline15_3 R8FR852819 right terminal Beta vulgaris subsp. Beta vulgaris repeatvulgarius subsp. vulgaris LINE-type retrotransposon Belline12_2 R8FR852813 right terminal Beta vulgaris subsp. Beta vulgaris repeatvulgarius subsp. vulgaris LINE-type retrotransposon Belline9_5 R8FR852807 right terminal Beta vulgaris subsp. Beta vulgaris repeatvulgarius subsp. vulgaris LINE-type retrotransposon Belline8_1 R8FR852806 right terminal Beta vulgaris subsp. Beta vulgaris repeatvulgarius subsp. vulgaris LINE-type retrotransposon Belline7_18 R8FR852799 right terminal Beta vulgaris subsp. Beta vulgaris repeatvulgarius subsp. vulgaris LINE-type retrotransposon Belline2_4 R8FR852795 right terminal Beta vulgaris subsp. Beta vulgaris repeatvulgarius subsp. vulgaris LINE-type retrotransposon BNR19 (Belline1_19)R8 AF352481 transposon: Chironomus circumdatus clone ChironomusNLRCth1-like cir8 transposon NLRCth1-like circumdatus non-LTR non-LTRretrotransposon retrotransposon reverse transcriptase gene, partial cdsR8; FR852861 right terminal Beta vulgaris subsp. Beta vulgaris R5 repeatvulgaris subsp. vulgaris LINE-type retrotransposon BNR59 (Belline1_59)R8; FR852857 right terminal Beta vulgaris subsp. Beta vulgaris R5 repeatvulgaris subsp. vulgaris LINE-type retrotransposon BNR51 (Belline1_51)R8; FR852866 right terminal Beta vulgaris subsp. Beta vulgaris R7 repeatvulgaris subsp. vulgaris LINE-type retrotransposon BNR76 (Belline1_76)R8; FR852838 right terminal Beta vulgaris subsp. Beta vulgaris R7 repeatvulgaris subsp. vulgaris LINE-type retrotransposon BNR7 (Belline1_7) R8;AF352455 transposon: Chironomus alpestris clone Chironomus R7NLRCth1-like dor70 note identical sequence alpestris non-LTR found indor80 transposon retrotransposon NLRCth1-like non-LTR retrotransposonreverse transcriptase gene, partial cds R8; FR852878 right terminal Betavulgaris subsp. Beta vulgaris R9 repeat vulgaris subsp. vulgarisLINE-type retrotransposon BNR96 (Belline1_96) R9 KJ958625 R2 Bacillusrossius non-LTR Bacillus rossius retrotransposon trKOR9, partialsequence Rex6 AJ293547 en Oreochromis niloticus Rex6 Oreochromisretrotransposon partial en niloticus pseudogene for endonuclease, clonerex6-Oni-3 Rex6 AJ293546 en Oreochromis niloticus Rex6 Oreochromisretrotransposon partial en niloticus pseudogene for endonuclease, clonerex6-Oni-2 Rex6 AJ293545 en Oreochromis niloticus Rex6 Oreochromisretrotransposon partial en niloticus pseudogene for endonuclease, clonerex6-Oni-1 Rex6 AJ293517 en Xiphophorus maculatus Rex6 Xiphophorusretrotransposon partial en maculatus pseudogene for endonuclease, cloneRex6-Xma-6 Rex6 AJ293516 en Xiphophorus maculatus Rex6 Xiphophorusretrotransposon partial en maculatus pseudogene for endonuclease, cloneRex6-Xma-5 Rex6 AJ293515 en Xiphophorus maculatus Rex6 Xiphophorusretrotransposon partial en maculatus pseudogene for endonuclease, cloneRex6-Xma-4 Rex6 AJ293514 en Xiphophorus maculatus Rex6 Xiphophorusretrotransposon partial en maculatus pseudogene for endonuclease, cloneRex6-Xma-3 Rex6 AJ293513 en Xiphophorus maculatus Rex6 Xiphophorusretrotransposon partial en maculatus pseudogene for endonuclease, cloneRex6-Xma-2 Rex6 AJ293512 en Xiphophorus maculatus Rex6 Xiphophorusretrotransposon partial en maculatus pseudogene for endonuclease, cloneRex6-Xma-1 Rex6 AJ293538 en Poecilia formosa Rex6 Poecilia formosaretrotransposon partial en pseudogene for endonuclease, clone rex6-Pfo-6Rex6 AJ293537 en Poecilia formosa Rex6 Poecilia formosa retrotransposonpartial en pseudogene for endonuclease, clone rex6-Pfo-5 Rex6 AJ293536en Poecilia formosa Rex6 Poecilia formosa retrotransposon partial enpseudogene for endonuclease, clone rex6-Pfo-4 Rex6 AJ293535 en Poeciliaformosa Rex6 Poecilia formosa retrotransposon partial en pseudogene forendonuclease, clone rex6-Pfo-3 Rex6 AJ293534 en Poecilia formosa Rex6Poecilia formosa retrotransposon partial en pseudogene for endonuclease,clone rex6-Pfo-2 Rex6 AJ293533 en Poecilia formosa Rex6 Poecilia formosaretrotransposon partial en pseudogene for endonuclease, clone rex6-Pfo-1Rex6 AJ293526 en Poeciliopsis gracilis Rex6 Poeciliopsis retrotransposonpartial en gracilis pseudogene for endonuclease, clone rex6-Pgr-4 Rex6AJ293525 en Poeciliopsis gracilis Rex6 Poeciliopsis retrotransposonpartial en gracilis pseudogene for endonuclease, clone rex6-Pgr-3 Rex6AJ293524 en Poeciliopsis gracilis Rex6 Poeciliopsis retrotransposonpartial en gracilis pseudogene for endonuclease, clone rex6-Pgr-2 Rex6AJ293523 en Poeciliopsis gracilis Rex6 Poeciliopsis retrotransposonpartial en gracilis pseudogene for endonuclease, clone rex6-Pgr-1 Rex6AJ293522 en Oryzias latipes Rex6 Oryzias latipes retrotransposon partialen pseudogene for endonuclease, clone rex6-Ola-5 Rex6 AJ293521 enOryzias latipes Rex6 Oryzias latipes retrotransposon partial enpseudogene for endonuclease, clone Rex6-Ola-4 Rex6 AJ293520 en Oryziaslatipes Rex6 Oryzias latipes retrotransposon partial en pseudogene forendonuclease, clone Rex6-Ola-3 Rex6 AJ293519 en Oryzias latipes Rex6Oryzias latipes retrotransposon partial en pseudogene for endonuclease,clone Rex6-Ola-2 Rex6 AJ293518 en Oryzias latipes Rex6 Oryzias latipesretrotransposon partial en pseudogene for endonuclease, clone Rex6-Ola-1Rex6 AJ293549 en Cichlasoma labridens Rex6 Herichthys retrotransposonpartial en labridens pseudogene for endonuclease, clone rex6-Cla-2 Rex6AJ293548 en Cichlasoma labridens Rex6 Herichthys retrotransposon partialen labridens pseudogene for endonuclease, clone rex6-Cla-1 Rex6 AJ293544en Heterandria bimaculata Rex6 Pseudoxiphophorus retrotransposon partialen bimaculatus pseudogene for endonuclease, clone rex6-Hbi-6 Rex6AJ293543 en Heterandria bimaculata Rex6 Pseudoxiphophorusretrotransposon partial en bimaculatus pseudogene for endonuclease,clone rex6-Hbi-5 Rex6 AJ293542 en Heterandria bimaculata Rex6Pseudoxiphophorus retrotransposon partial en bimaculatus pseudogene forendonuclease, clone rex6-Hbi-4 Rex6 AJ293541 en Heterandria bimaculataRex6 Pseudoxiphophorus retrotransposon partial en bimaculatus pseudogenefor endonuclease, clone rex6-Hbi-3 Rex6 AJ293540 en Heterandriabimaculata Rex6 Pseudoxiphophorus retrotransposon partial en bimaculatuspseudogene for endonuclease, clone rex6-Hbi-2 Rex6 AJ293539 enHeterandria bimaculata Rex6 Pseudoxiphophorus retrotransposon partial enbimaculatus pseudogene for endonuclease, clone rex6-Hbi-1 Rex6 AJ293532en Gambusia affinis Rex6 Gambusia affinis retrotransposon partial enpseudogene for endonuclease, clone rex6-Gaf-5 Rex6 AJ293531 en Gambusiaaffinis Rex6 Gambusia affinis retrotransposon partial en pseudogene forendonuclease, clone rex6-Gaf-5 Rex6 AJ293530 en Gambusia affinis Rex6Gambusia affinis retrotransposon partial en pseudogene for endonuclease,clone rex6-Gaf-4 Rex6 AJ293529 en Gambusia affinis Rex6 Gambusia affinisretrotransposon partial en pseudogene for endonuclease, clone rex6-Gaf-3Rex6 AJ293528 en Gambusia affinis Rex6 Gambusia affinis retrotransposonpartial en pseudogene for endonuclease, clone rex6-Gaf-2 Rex6 AJ293527en Gambusia affinis Rex6 Gambusia affinis retrotransposon partial enpseudogene for endonuclease, clone rex6-Gaf-1 Rex6 JX576459 non-LTRSymphysodon discus isolate Symphysodon discus retrotransposon: i non-LTRretrotransposon Rex6 Rex6, partial sequence Rex6 JX576458 non-LTRSymphysodon discus isolate Symphysodon discus retrotransposon: h non-LTRretrotransposon Rex6 Rex6, partial sequence Rex6 JX576457 non-LTRSymphysodon discus isolate Symphysodon discus retrotransposon: g non-LTRretrotransposon Rex6 Rex6, partial sequence Rex6 JX576456 non-LTRSymphysodon discus isolate Symphysodon discus retrotransposon: f non-LTRretrotransposon Rex6 Rex6, partial sequence Rex6 JX576455 non-LTRSymphysodon discus isolate Symphysodon discus retrotransposon: e non-LTRretrotransposon Rex6 Rex6, partial sequence Rex6 JX576454 non-LTRSymphysodon discus isolate Symphysodon discus retrotransposon: d non-LTRretrotransposon Rex6 Rex6, partial sequence Rex6 JX576453 non-LTRSymphysodon discus isolate Symphysodon discus retrotransposon: c non-LTRretrotransposon Rex6 Rex6, partial sequence Rex6 JX576452 non-LTRSymphysodon discus isolate Symphysodon discus retrotransposon: b non-LTRretrotransposon Rex6 Rex6, partial sequence Rex6 JX576451 non-LTRSymphysodon discus isolate Symphysodon discus retrotransposon: a non-LTRretrotransposon Rex6 Rex6, partial sequence Rex6 JX576450 non-LTRSymphysodon discus isolate Symphysodon discus retrotransposon: z8non-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576449non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon:z7 non-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576448non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon:z6 non-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576447non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon:z5 non-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576446non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon:z4 non-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576445non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon:z3 non-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576444non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon:z2 non-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576443non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon:z1 non-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576442non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon: znon-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576441non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon: xnon-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576440non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon: vnon-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576439non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon: unon-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576438non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon: tnon-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576437non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon: snon-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576436non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon: rnon-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576435non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon: qnon-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576434non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon: pnon-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576433non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon: onon-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576432non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon: nnon-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576431non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon: mnon-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576430non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon: lnon-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576429non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon: knon-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576428non-LTR Symphysodon discus isolate Symphysodon discus retrotransposon: jnon-LTR retrotransposon Rex6 Rex6, partial sequence Rex6 JX576427non-LTR Pterophyllum scalare clone Pterophyllum retrotransposon: enon-LTR retrotransposon scalare Rex6 Rex6, partial sequence Rex6JX576426 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: d non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 JX576425 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: c non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 JX576424 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: b non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 JX576423 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: a non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 JX576422 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: g non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 JX576421 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: f non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 JX576420 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: e non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 JX576419 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: d non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 JX576418 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: c non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 JX576417 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: b non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 JX576416 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: a non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 JX576415 non-LTR Astronotus ocellatus clone Astronotusretrotransposon: g non-LTR retrotransposon ocellatus Rex6 Rex6, partialsequence Rex6 JX576414 non-LTR Astronotus ocellatus clone Astronotusretrotransposon: f non-LTR retrotransposon ocellatus Rex6 Rex6, partialsequence Rex6 JX576413 non-LTR Astronotus ocellatus clone Astronotusretrotransposon: e non-LTR retrotransposon ocellatus Rex6 Rex6, partialsequence Rex6 JX576412 non-LTR Astronotus ocellatus clone Astronotusretrotransposon: d non-LTR retrotransposon ocellatus Rex6 Rex6, partialsequence Rex6 JX576411 non-LTR Astronotus ocellatus clone Astronotusretrotransposon: c non-LTR retrotransposon ocellatus Rex6 Rex6, partialsequence Rex6 JX576410 non-LTR Astronotus ocellatus clone Astronotusretrotransposon: b non-LTR retrotransposon ocellatus Rex6 Rex6, partialsequence Rex6 JX576409 non-LTR Astronotus ocellatus clone Astronotusretrotransposon: a non-LTR retrotransposon ocellatus Rex6 Rex6, partialsequence Rex6 JX576408 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: h non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 JX576407 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: g non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 JX576406 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: f non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 JX576405 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: e non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 JX576404 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: d non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 JX576403 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: c non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 JX576402 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: b non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 JX576401 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: a non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 KF131853 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: z7 non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131852 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: z6 non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131851 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: z5 non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131850 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: z4 non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131849 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: z3 non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131848 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: z2 non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131847 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: z1 non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131846 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: z non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131845 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: x non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131844 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: v non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131843 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: u non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131842 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: t non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131841 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: s non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131840 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: r non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131839 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: q non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131838 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: p non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131837 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: n non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131836 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: m non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131835 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: l non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131834 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: k non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131833 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: j non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131832 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: i non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131831 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: h non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131830 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: g non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131829 non-LTR Pterophyllum scalare clone Pterophyllumretrotransposon: f non-LTR retrotransposon scalare Rex6 Rex6, partialsequence Rex6 KF131828 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: z6 non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131827 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: z5 non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131826 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: z4 non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131825 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: z3 non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131824 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: z2 non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131823 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: z1 non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131822 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: z non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131821 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: x non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131820 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: v non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131819 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: u non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131818 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: t non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131817 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: s non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131816 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: r non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131815 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: q non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131814 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: p non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131813 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: o non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131812 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: n non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131811 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: m non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131810 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: l non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131809 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: k non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131808 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: j non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131807 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: i non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131806 non-LTR Geophagus proximus clone Geophagusproximus retrotransposon: h non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131805 non-LTR Astronotus ocellatus clone Astronotusretrotransposon: z10 non-LTR retrotransposon ocellatus Rex6 Rex6,partial sequence Rex6 KF131804 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: z9 non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131803 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: z8 non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131802 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: z7 non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131801 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: z6 non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131800 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: z5 non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131799 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: z4 non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131798 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: z3 non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131797 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: z2 non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131796 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: z1 non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131795 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: z non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131794 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: x non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131793 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: v non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131792 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: u non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131791 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: t non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131790 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: s non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131789 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: r non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131788 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: q non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131787 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: p non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131786 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: o non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131785 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: n non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131784 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: m non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131783 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: l non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131782 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: k non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131781 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: j non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131780 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: i non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131779 non-LTR Astronotus ocellatus cloneAstronotus retrotransposon: h non-LTR retrotransposon ocellatus Rex6Rex6, partial sequence Rex6 KF131778 non-LTR Cichla monoculus cloneCichla monoculus retrotransposon: z6 non-LTR retrotransposon Rex6 Rex6,partial sequence Rex6 KF131777 non-LTR Cichla monoculus clone Cichlamonoculus retrotransposon: z5 non-LTR retrotransposon Rex6 Rex6, partialsequence Rex6 KF131776 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: z4 non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 KF131775 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: z3 non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 KF131774 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: z2 non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 KF131773 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: z1 non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 KF131772 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: z non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 KF131771 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: x non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 KF131770 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: v non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 KF131769 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: u non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 KF131768 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: t non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 KF131767 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: s non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 KF131766 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: r non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 KF131765 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: q non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 KF131764 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: p non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 KF131763 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: o non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 KF131762 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: n non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 KF131761 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: m non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 KF131760 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: l non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 KF131759 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: k non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 KF131758 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: j non-LTR retrotransposon Rex6 Rex6, partial sequenceRex6 KF131757 non-LTR Cichla monoculus clone Cichla monoculusretrotransposon: i non-LTR retrotransposon Rex6 Rex6, partial sequenceSLACS JN608782 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: Y-46 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608781 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: Y-45 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608780 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: Y-41 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608779 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: Y-40 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608778non-LTR Silene latifolia isolate Silene latifolia retrotransposon: Y-38non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608777 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: Y-37 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608776 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: Y-36 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608775 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: Y-35 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608774 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: Y-34 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608773non-LTR Silene latifolia isolate Silene latifolia retrotransposon: Y-33non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608772 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: Y-32 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608771 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: Y-30 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608770 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: Y-29 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608769 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: Y-28 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608768non-LTR Silene latifolia isolate Silene latifolia retrotransposon: Y-27non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608767 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: Y-26 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608766 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: Y-25 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608765 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: Y-24 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608764 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: Y-23 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608763non-LTR Silene latifolia isolate Silene latifolia retrotransposon: Y-22non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608762 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: Y-20 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608761 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: Y-19 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608760 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: Y-18 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608759 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: Y-17 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608758non-LTR Silene latifolia isolate Silene latifolia retrotransposon: Y-16non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608757 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: Y-13 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608756 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: Y-12 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608755 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: Y-11 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608754 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: Y-10 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608753non-LTR Silene latifolia isolate Silene latifolia retrotransposon: Y-08non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608752 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: Y-07 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608751 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: Y-06 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608750 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: Y-05 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608749 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: Y-04 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608748non-LTR Silene latifolia isolate Silene latifolia retrotransposon: Y-03non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608747 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: Y-01 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608746 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: X-83 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608745 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: X-81 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608744 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: X-80 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608743non-LTR Silene latifolia isolate Silene latifolia retrotransposon: X-79non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608742 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: X-78 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608741 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: X-76 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608740 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: X-75 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608739 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: X-74 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608738non-LTR Silene latifolia isolate Silene latifolia retrotransposon: X-66non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608737 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: X-65 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608736 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: X-62 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608735 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: X-61 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608734 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: X-60 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608733non-LTR Silene latifolia isolate Silene latifolia retrotransposon: X-59non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608732 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: X-57 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608731 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: X-49 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608730 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: X-41 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608729 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: X-28 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608728non-LTR Silene latifolia isolate Silene latifolia retrotransposon: X-27non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608727 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: X-23 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608726 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: X-21 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608725 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: X-14 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608724 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: X-11 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608723non-LTR Silene latifolia isolate Silene latifolia retrotransposon: X-07non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608722 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: X-06 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608721 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: X-05 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608720 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: X-03 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608719 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: X-01 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608718non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mG40non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608717 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mG39 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608716 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mG38 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608715 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mG37 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608714 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mG36 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608713non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mG35non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608712 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mG34 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608711 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mG33 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608710 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mG32 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608709 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mG31 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608708non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mG30non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608707 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mG29 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608706 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mG28 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608705 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mG27 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608704 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mG26 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608703non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mG25non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608702 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mG24 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608701 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mG23 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608700 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mG22 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608699 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mG21 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608698non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mG20non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608697 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mG19 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608696 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mG18 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608695 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mG17 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608694 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mG16 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608693non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mG14non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608692 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mG13 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608691 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mG12 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608690 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mG11 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608689 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mG10 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608688non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mG09non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608687 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mG08 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608686 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mG07 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608685 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mG06 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608684 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mG05 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608683non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mG04non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608682 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mG03 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608681 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mG02 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608680 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mG01 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608679 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fG40 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608678non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fG39non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608677 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fG38 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608676 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fG37 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608675 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fG36 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608674 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fG35 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608673non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fG34non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608672 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fG33 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608671 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fG32 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608670 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fG31 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608669 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fG30 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608668non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fG29non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608667 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fG28 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608666 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fG27 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608665 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fG26 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608664 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fG25 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608663non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fG24non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608662 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fG23 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608661 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fG22 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608660 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fG21 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608659 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fG20 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608658non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fG19non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608657 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fG18 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608656 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fG17 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608655 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fG16 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608654 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fG15 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608653non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fG14non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608652 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fG13 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608651 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fG11 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608650 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fG10 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608649 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fG09 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608648non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fG08non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608647 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fG07 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608646 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fG06 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608645 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fG05 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608644 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fG03 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608643non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fG02non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608642 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fG01 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608641 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: A-47 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608640 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: A-46 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608639 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: A-45 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608638non-LTR Silene latifolia isolate Silene latifolia retrotransposon: A-44non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608637 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: A-42 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608636 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: A-41 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608635 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: A-40 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608634 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: A-37 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608633non-LTR Silene latifolia isolate Silene latifolia retrotransposon: A-36non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608632 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: A-35 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608631 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: A-34 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608630 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: A-33 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608629 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: A-32 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608628non-LTR Silene latifolia isolate Silene latifolia retrotransposon: A-31non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608627 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: A-26 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608626 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: A-24 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608625 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: A-23 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608624 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: A-22 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608623non-LTR Silene latifolia isolate Silene latifolia retrotransposon: A-21non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608622 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: A-20 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608621 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: A-19 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608620 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: A-18 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608619 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: A-17 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608618non-LTR Silene latifolia isolate Silene latifolia retrotransposon: A-16non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608617 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: A-15 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608616 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: A-14 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608615 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: A-13 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608614 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: A-12 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608613non-LTR Silene latifolia isolate Silene latifolia retrotransposon: A-11non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608612 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: A-10 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608611 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: A-09 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608610 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: A-08 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608609 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: A-06 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608608non-LTR Silene latifolia isolate Silene latifolia retrotransposon: A-05non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608607 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: A-03 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608606 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: A-01 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608236 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mR40 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608235 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mR39 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608234non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mR38non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608233 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mR35 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608232 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mR32 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608231 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mR29 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608230 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mR26 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608229non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mR25non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608228 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mR18 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608227 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mR17 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608226 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mR14 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608225 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mR13 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608224non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mR12non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608223 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mR11 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608222 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mR09 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608221 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mR08 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608220 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mR07 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608219non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mR06non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608218 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mR05 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608217 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mR04 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608216 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mR03 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608215 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mR02 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608214non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mR01non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608213 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mL40 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608212 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mL39 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608211 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mL37 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608210 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mL36 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608209non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mL35non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608208 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mL34 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608207 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mL33 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608206 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mL32 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608205 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mL31 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608204non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mL28non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608203 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mL27 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608202 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mL25 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608201 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mL24 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608200 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mL23 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608199non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mL19non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608198 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mL16 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608197 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mL13 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608196 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mL12 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608195 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mL11 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608194non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mL10non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608193 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mL07 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608192 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mL04 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608191 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mL03 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608190 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mL02 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608189non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mL01non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608188 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mF40 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608187 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mF39 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608186 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mF38 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608185 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mF37 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608184non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mF36non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608183 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mF35 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608182 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mF33 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608181 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mF32 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608180 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mF31 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608179non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mF30non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608178 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mF29 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608177 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mF28 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608176 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mF27 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608175 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mF26 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608174non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mF25non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608173 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mF23 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608172 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mF22 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608171 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mF21 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608170 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mF20 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608169non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mF19non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608168 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mF18 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608167 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mF17 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608166 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mF16 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608165 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mF15 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608164non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mF14non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608163 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mF13 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608162 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mF11 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608161 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mF10 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608160 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mF09 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608159non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mF08non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608158 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mF07 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608157 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mF06 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608156 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: mF05 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608155 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: mF04 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608154non-LTR Silene latifolia isolate Silene latifolia retrotransposon: mF03non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608153 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: mF02 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608152 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: mF01 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608151 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fR40 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608150 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fR39 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608149non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fR38non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608148 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fR37 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608147 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fR36 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608146 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fR35 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608145 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fR34 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608144non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fR33non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608143 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fR32 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608142 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fR31 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608141 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fR30 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608140 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fR29 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608139non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fR28non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608138 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fR27 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608137 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fR26 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608136 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fR25 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608135 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fR24 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608134non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fR23non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608133 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fR22 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608132 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fR21 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608131 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fR20 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608130 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fR19 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608129non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fR18non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608128 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fR17 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608127 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fR16 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608126 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fR15 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608125 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fR14 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608124non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fR13non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608123 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fR12 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608122 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fR11 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608121 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fR09 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608120 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fR08 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608119non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fR07non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608118 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fR05 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608117 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fR04 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608116 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fR03 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608115 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fR02 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608114non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fR01non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608113 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fL40 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608112 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fL39 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608111 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fL38 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608110 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fL37 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608109non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fL36non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608108 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fL35 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608107 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fL34 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608106 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fL33 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608105 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fL32 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608104non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fL31non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608103 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fL30 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608102 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fL29 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608101 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fL28 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608100 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fL27 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608099non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fL26non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608098 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fL25 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608097 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fL24 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608096 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fL23 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608095 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fL22 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608094non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fL20non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608093 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fL19 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608092 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fL18 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608091 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fL17 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608090 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fL16 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608089non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fL15non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608088 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fL14 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608087 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fL13 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608086 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fL12 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608085 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fL11 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608084non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fL10non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608083 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fL09 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608082 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fL07 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608081 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fL06 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608080 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fL05 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608079non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fL04non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608078 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fL03 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608077 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fL01 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608076 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fF40 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608075 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fF39 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608074non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fF38non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608073 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fF37 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608072 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fF36 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608071 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fF35 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608070 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fF34 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608069non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fF33non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608068 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fF32 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608067 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fF31 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608066 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fF30 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608065 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fF29 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608064non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fF28non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608063 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fF27 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608062 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fF26 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608061 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fF24 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608060 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fF23 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608059non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fF21non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608058 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fF20 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608057 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fF18 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608056 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fF17 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608055 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fF16 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608054non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fF15non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608053 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fF14 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608052 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fF13 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608051 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fF12 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608050 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fF11 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608049non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fF10non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608048 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fF09 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608047 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fF08 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence SLACS JN608046 non-LTR Silene latifoliaisolate Silene latifolia retrotransposon: fF06 non-LTR retrotransposonSLACS-like SLACS-like, partial sequence SLACS JN608045 non-LTR Silenelatifolia isolate Silene latifolia retrotransposon: fF05 non-LTRretrotransposon SLACS-like SLACS-like, partial sequence SLACS JN608044non-LTR Silene latifolia isolate Silene latifolia retrotransposon: fF04non-LTR retrotransposon SLACS-like SLACS-like, partial sequence SLACSJN608043 non-LTR Silene latifolia isolate Silene latifoliaretrotransposon: fF03 non-LTR retrotransposon SLACS-like SLACS-like,partial sequence SLACS JN608042 non-LTR Silene latifolia isolate Silenelatifolia retrotransposon: fF02 non-LTR retrotransposon SLACS-likeSLACS-like, partial sequence YURECi AB097133 rt Ciona intestinalis Cionaintestinalis retrotransposon YURECi DNA, complete sequence CRE . Cnl1 C.neoformans non-LTR Cryptococcus retrotransposon - consensus. neoformansCRE . CRE-1_ACas CRE non-LTR retrotransposon: Acanthamoeba consensus.castellanii CRE . Cre-1_BM Cre-1_BM non-LTR Bombyx moriretrotransposon - consensus. CRE . CRE-1_CCri Non-LTR retrotransposonfrom Chondrus crispus the red seaweed: consensus. CRE . Cre-1_FCyCre-1_FCy non-LTR Fragilariopsis retrotransposon - conceptual cylindrusconsensus. CRE . Cre-1_HM Cre-1_HM non-LTR Hydra vulgarisretrotransposon - consensus. CRE . CRE-1_HRo Cre-like non-LTR Helobdellarobusta retrotransposon: consensus sequence. CRE . CRE-1_LSa CRE non-LTRretrotransposon: Lactuca sativa consensus. CRE . Cre-1_MB Cre-1_MBnon-LTR Monosiga retrotransposon - consensus. brevicollis CRE . Cre-1_NVCre-1_NV non-LTR Nematostella retrotransposon - consensus. vectensis CRE. CRE-1_PXu Non-LTR retrotransposon from Papilio xuthus Papilio xuthus:consensus. CRE . CRE-10_CCri Non-LTR retrotransposon from Chondruscrispus the red seaweed: consensus. CRE . CRE-11_CCri Non-LTRretrotransposon from Chondrus crispus the red seaweed: consensus. CRE .CRE-12_CCri Non-LTR retrotransposon from Chondrus crispus the redseaweed: consensus. CRE . CRE-13_CCri Non-LTR retrotransposon fromChondrus crispus the red seaweed: consensus. CRE . CRE-14_CCri Non-LTRretrotransposon from Chondrus crispus the red seaweed: consensus. CRE .CRE-15_CCri Non-LTR retrotransposon from Chondrus crispus the redseaweed: consensus. CRE . CRE-16_CCri Non-LTR retrotransposon fromChondrus crispus the red seaweed: consensus. CRE . CRE-17_CCri Non-LTRretrotransposon from Chondrus crispus the red seaweed: consensus. CRE .Cre-2_BM Cre-2_BM non-LTR Bombyx mori retrotransposon - consensus. CRE .CRE-2_CCri Non-LTR retrotransposon from Chondrus crispus the redseaweed: consensus. CRE . CRE-2_HMa CRE non-LTR retrotransposon: Hydravulgaris consensus. CRE . CRE-2_HRo Cre-like non-LTR Helobdella robustaretrotransposon: consensus sequence. CRE . CRE-2_NV CRE non-LTRretrotransposon: Nematostella consensus. vectensis CRE . CRE-2_PXuNon-LTR retrotransposon from Papilio xuthus Papilio xuthus: consensus.CRE . CRE-3_CCri Non-LTR retrotransposon from Chondrus crispus the redseaweed: consensus. CRE . CRE-3_HRo CRE-like non-LTR Helobdella robustaretrotransposon: consensus sequence. CRE . CRE-3_NV CRE non-LTRretrotransposon: Nematostella consensus. vectensis CRE . CRE-4_CCriNon-LTR retrotransposon from Chondrus crispus the red seaweed:consensus. CRE . CRE-4_HRo CRE-like non-LTR Helobdella robustaretrotransposon: consensus sequence. CRE . CRE-5_CCri Non-LTRretrotransposon from Chondrus crispus the red seaweed: consensus. CRE .CRE-5_HRo CRE-like non-LTR Helobdella robusta retrotransposon: consensussequence. CRE . CRE-6_CCri Non-LTR retrotransposon from Chondrus crispusthe red seaweed: consensus. CRE . CRE-6_HRo CRE-like non-LTR Helobdellarobusta retrotransposon: consensus sequence. CRE . CRE-7_CCri Non-LTRretrotransposon from Chondrus crispus the red seaweed: consensus. CRE .CRE-8_CCri Non-LTR retrotransposon from Chondrus crispus the redseaweed: consensus. CRE . CRE-9_CCri Non-LTR retrotransposon fromChondrus crispus the red seaweed: consensus. CRE M33009 CRE1 C.fasciculata retrotransposable Crithidia element (CRE1). fasciculata CREU19151 CRE2 C. fasciculata retrotransposable Crithidia element (CRE2).fasciculata CRE M62862 CZAR T. cruzi SL-RNA-associated Trypanosoma cruzinon-LTR retrotransposon. R4 . Dong Bombyx mori non-LTR Bombyx moriretrotransposable element. R4 . DONG_FR2 Non-LTR retrotransposon;Takifugu rubripes site-specific LINE; R4/Dong superfamily; DONG_FR2. R4. Dong-1_AFC Dong/R4-type non-LTR Cichlidae retrotransposon - consensus.R4 . Dong-1_HMM Non-LTR retrotransposon Heliconius family fromHeliconius melpomene melpomene melpomene. melpomene R4 . Dong-1_NVe ADong non-LTR Nematostella retrotransposon family from vectensisNematostella vectensis. R4 . Dong-1_PPo Non-LTR retrotransposon fromPapilio polytes Papilio polytes: consensus. R4 . Dong-1_PXu Non-LTRretrotransposon from Papilio xuthus Papilio xuthus: consensus. R4 .Dong-2_BM Non-LTR retrotransposon - a Bombyx mori consensus. R4 .Dong-2_HMM Non-LTR retrotransposon Heliconius family from Heliconiusmelpomene melpomene melpomene. melpomene R4 . Dong-2_Lch Dong-likenon-LTR Latimeria retrotransposon - consensus. chalumnae R4 . Dong-2_PPoNon-LTR retrotransposon from Papilio polytes Papilio polytes: consensus.R4 . DongAa A Dong non-LTR Aedes aegypti retrotransposon family fromAedes aegypti. R4 AB097127 DongAG Anopheles gambiae non-LTR Anophelesretrotransposon DongAg - a gambiae partial sequence. R4 AB097128 EhRLE2Entamoeba histolytica Entamoeba retrotransposon EhRLE2, histolyticacomplete sequence. R4 AB097129 EhRLE3 Entamoeba histolytica Entamoebaretrotransposon EhRLE3, histolytica complete sequence. HERO . HERO-1_AFCHero-type non-LTR Cichlidae retrotransposon - consensus. HERO .HERO-1_BF Amphioxus HERO-1_BF Branchiostoma autonomous non-LTR floridaeRetrotransposon - consensus. HERO . HERO-1_HR A family of HERO non-LTRHelobdella robusta retrotransposons - a consensus sequence. HERO .HERO-1_PP A family of HERO non-LTR Physarum retrotransposons - apolycephalm consensus sequence. HERO AAGJ02121261 HERO-1_SP Sea urchinHERO-1_SP Strongylocentrotus autonomous non-LTR purpuratusRetrotransposon - consensus. HERO . HERO-2_BF Amphioxus HERO-2_BFBranchiostoma autonomous non-LTR floridae Retrotransposon - consensus.HERO . HERO-2_DR HERO-2_DR is a family of HERO Danio rerio non-LTRretrotransposons - a consensus. HERO . HERO-2_HR Non-LTRretrotransposon: Helobdella robusta consensus sequence. HERO 048B05Hero-2_SPur HERO-type non-ltr Strongylocentrotus retrotransposon fromsea urchin. purpuratus HERO . HERO-3_BF HERO-3_BF is a family of HEROBranchiostoma non-LTR retrotransposons - a floridae consensus. HERO .HERO-3_DR HERO-3_DR is a family of HERO Danio rerio non-LTRretrotransposons - a consensus. HERO . HERO-3_HR Non-LTRretrotransposon: Helobdella robusta consensus sequence. HERO .Hero-3_SPur HERO-type non-LTR Strongylocentrotus retrotransposon fromsea urchin. purpuratus HERO . HERO-4_DR HERO-4_DR is a family of HERODanio rerio non-LTR retrotransposons - a consensus. HERO . HERO-4_HRNon-LTR retrotransposon: Helobdella robusta consensus sequence. HERO .HERO-5_HR Non-LTR retrotransposon: Helobdella robusta consensussequence. HERO . HERO-6_HR Non-LTR retrotransposon: Helobdella robustaconsensus sequence. HERO . HERO-7_HR Non-LTR retrotransposon: Helobdellarobusta consensus sequence. HERO . HERO-8_HR Non-LTR retrotransposon:Helobdella robusta consensus sequence. HERO . HERO-9_HR Non-LTRretrotransposon: Helobdella robusta consensus sequence. HERO . HERODrHERODr is a family of HERO Danio rerio non-LTR retrotransposons - aconsensus. HERO . HEROFr A HERO clade non-LTR Takifugu rubripesRetrotransposon family - consensus. HERO . HEROTn HEROTn or Zebulonnon-LTR Tetraodon retrotransposon - a consensus nigroviridis sequence.NeSL . LIN10B_SM Non-LTR retrotransposon; Schmidtea consensus.mediterranea NeSL . LIN11_SM Non-LTR retrotransposon: Schmidteaconsensus. mediterranea NeSL . LIN13_SM Non-LTR retrotransposon;Schmidtea consensus. mediterranea NeSL . LIN14_SM Non-LTRretrotransposon; Schmidtea consensus. mediterranea NeSL . LIN15_SMNon-LTR retrotransposon; Schmidtea consensus. mediterranea NeSL .LIN2_SM Non-LTR retrotransposon Schmidtea (consensus). mediterranea NeSL. LIN21_SM Non-LTR retrotransposon; Schmidtea consensus. mediterraneaNeSL . LIN23_SM Non-LTR retrotransposon; Schmidtea consensus.mediterranea NeSL . LIN24_SM Non-LTR retrotransposon; Schmidteaconsensus. mediterranea NeSL . LIN24B_SM Non-LTR retrotransposon;Schmidtea consensus. mediterranea NeSL . LIN25_SM Non-LTRretrotransposon; Schmidtea consensus. mediterranea NeSL . LIN26_SMNon-LTR retrotransposon; Schmidtea consensus. mediterranea NeSL .LIN3_SM Non-LTR retrotransposon; Schmidtea consensus. mediterranea NeSL. LIN4_SM Non-LTR retrotransposon; Schmidtea consensus. mediterraneaNeSL . LIN4b_SM Non-LTR retrotransposon; Schmidtea consensus.mediterranea NeSL . LIN5_SM Non-LTR retrotransposon from SchmidteaSchmidtea mediterranea: mediterranea consensus. NeSL . LIN6_SM Non-LTRretrotransposon from Schmidtea Schmidtea mediterranea: mediterraneaconsensus. NeSL . LIN7_SM Non-LTR retrotransposon from SchmidteaSchmidtea mediterranea: mediterranea consensus. NeSL . LIN7B_SM Non-LTRretrotransposon; Schmidtea consensus. mediterranea NeSL . LIN9_SMNon-LTR retrotransposon: Schmidtea consensus. mediterranea CRE JQ747487MoTeR1 Telomere-specific non-LTR Magnaporthe oryzae retrotransposonMoTeR1 from Magnaporthe oryzae. CRE JQ747488 MoTeR2 Telomere-specificnon-LTR Magnaporthe oryzae retrotransposon MoTeR2 from Magnaportheoryzae. NeSL Z82058 NeSL-1 NeSL-1 is a non-LTR Caenorhabditisretrotransposon, complete elegans sequence. NeSL . NeSL-1_C11 A familyof NeSL non-LTR Caenorhabditis retrotransposons. tropicalis NeSL .NeSL-1_CA A family of NeSL non-LTR Caenorhabditis retrotransposons.angaria NeSL . NeSL-1_CBre A family of NeSL non-LTR Caenorhabditisretrotransposons - consensus. brenneri NeSL . NeSL-1_CBri A family ofNeSL non-LTR Caenorhabditis retrotransposons. briggsae NeSL .NeSL-1_CJap A family of NeSL non-LTR Caenorhabditis retrotransposons -consensus. japonica NeSL . NeSL-1_CRem A family of NeSL non-LTRCaenorhabditis retrotransposons - consensus. remanei NeSL . NeSL-1_SMNon-LTR retrotransposon; Schmidtea consensus. mediterranea NeSL .NeSL-1_TV A family of NeSL non-LTR Trichomonas retrotransposons -consensus. vaginalis NeSL . NeSL-2_CBre A family of NeSL non-LTRCaenorhabditis retrotransposons - consensus. brenneri NeSL . NeSL-2_CRemA family of NeSL non-LTR Caenorhabditis retrotransposons - consensus.remanei NeSL . NeSL-2_SM Non-LTR retrotransposon; Schmidtea consensus.mediterranea NeSL . NeSL-3_CBre A family of NeSL non-LTR Caenorhabditisretrotransposons - consensus. brenneri NeSL . NeSL-3_CRem A family ofNeSL non-LTR Caenorhabditis retrotransposons - consensus. remanei NeSLchrUn NeSL-4_CRem A family of NeSL non-LTR Caenorhabditisretrotransposons. remanei NeSL . NeSL-4_SM Non-LTR retrotransposon;Schmidtea consensus. mediterranea R2 BN000800 PERERE-9 Schistosomamansoni Perere-9 Schistosoma mansoni non-LTR retrotransposon (EST). R4 .Plat_R4 R4 Non-LTR Retrotransposon Ornithorhynchus from Ornithorhynchus.R2 AF015815 R2_AM Anurida maritima Anurida maritima retrotransposon R2,complete sequence. R2 M16558 R2_BM Bombyx mori rDNA insertion Bombyxmori element R2 (type II), complete cds. R2 AB097121 R2CI R2-type LINE.Ciona intestinalis R2 . R2CPB Non-LTR retrotransposon: Chrysemyspictaconsensus. bellii R2 . R2_DAn 28S rDNA-specific non-LTR Drosophilaretrotransposon R2 in ananassae Drosophila ananassae. R2 X51967 R2_DMLINE-like retrotransposable Drosophila element R2DM. melanogaster R2 .R2_DPe 28S rDNA-specific non-LTR Drosophila retrotransposon R2 inpersimilis Drosophila persimilis. R2 . R2_DPs 28S rDNA-specific non-LTRDrosophila retrotransposon R2 in pseudoobscura Drosophila pseudoobscura.R2 . R2_DSe 28S rDNA-specific non-LTR Drosophila retrotransposon R2 insechellia Drosophila sechellia. R2 . R2_DSi 28S rDNA-specific non-LTRDrosophila retrotransposon R2 in simulans Drosophila simulans. R2 .R2_DYa 28S rDNA-specific non-LTR Drosophila yakuba retrotransposon R2 inDrosophila yakuba. R2 AF015819 R2_FA Forficula auricularia Forficularetrotransposon R2, complete auricularia sequence. R2 AF015816 R2_HCHippodamia convergens Hippodamia retrotransposon R2 reverse convergenstranscriptase gene, partial cds. R2 GU949558 R2_KF 28S rDNA-specificnon-LTR Kalotermes retrotransposon R2 from flavicollis Kalotermesflavicollis. R2 AF015814 R2_LP Limulus polyphemus Limulus polyphemusretrotransposon R2, complete sequence. R2 AF015818 R2_PS Porcellioscaber Porcellio scaber retrotransposon R2, complete sequence. R2GU949555 R2_RL 28S rDNA-specific non-LTR Reticulitermes retrotransposonR2 from lucifugus Reticulitermes lucifugus. R2 GU949554 R2_RU 28SrDNA-specific non-LTR Reticulitermes retrotransposon R2 from urbisReticulitermes urbis. R2 . R2-1_AAm R2 non-LTR retrotransposon Amblyommafrom lone star tick. americanum R2 . R2-1_ACC R2 non-LTR retrotransposonAquila chrysaetos from golden eagle. canadensis R2 . R2-1_ACh R2 non-LTRretrotransposon Acanthisitta from rifleman. chloris R2 . R2-1_AFo R2non-LTR retrotransposon Aptenodytes from emperor penguin. forsteri R2 .R2-1_AMi R2-type non-LTR retrotransposon. Alligator mississippiensis R2. R2-1_AOM R2 non-LTR retrotransposon Apteryx spp. from kiwi. R2 .R2-1_ApA R2 non-LTR retrotransposon Apteryx australis from north islandbrown kiwi. mantelli R2 . R2-1_APi R2 non-LTR retrotransposonAcyrthosiphon from pea aphid. pisum R2 . R2-1_BRG R2 non-LTRretrotransposon Balearica from East African grey regulorum crownedcrane. gibbericeps R2 . R2-1_BTe R2 non-LTR retrotransposon Bombusterrestris from buff-tailed bumblebee. R2 . R2-1_CAnn R2 non-LTRretrotransposon Calypte anna from Anna's hummingbird. R2 . R2-1_CAu R2non-LTR retrotransposon Cathartes aura from turkey vulture. R2 .R2-1_CBr R2 non-LTR retrotransposon Corvus from American crow.brachyrhynchos R2 . R2-1_CCa R2 non-LTR retrotransposon Antrostomus fromchuck-will's-widow. carolinensis R2 . R2-1_CCan R2 non-LTRretrotransposon Cuculus canorus from common cuckoo. R2 . R2-1_CPu R2non-LTR retrotransposon Calidris pugnax from ruff. R2 . R2-1_Crp Non-LTRretrotransposon. Crocodylus porosus R2 . R2-1_CSt R2 non-LTRretrotransposon Colius striatus from speckled mousebird. R2 . R2-1_CU R2non-LTR retrotransposon Chlamydotis from MacQueen's bustard. macqueeniiR2 . R2-1_CVo R2 non-LTR retrotransposon Charadrius from killdeer.vociferus R2 . R2-1_DWi 28S rDNA-specific non-LTR Drosophilaretrotransposon R2 in willistoni Drosophila willistoni. R2 . R2-1_EGa R2non-LTR retrotransposon Egretta garzetta from little egret. R2 .R2-1_FAl R2 non-LTR retrotransposon Ficedula from collared flycatcher.albicollis R2 . R2-1_FCh R2 non-LTR retrotransposon Falco cherrug fromSaker falcon. R2 . R2-1_FPe R2 non-LTR retrotransposon Falco peregrinusfrom peregrine falcon. R2 . R2-1_GA R2 non-LTR retrotransposonGasterosteus from three-spined stickleback. aculeatus R2 . R2-1_GavNon-LTR retrotransposon. Gavialis gangeticus R2 . R2-1_GFo R2 non-LTRretrotransposon Geospiza fortis from medium ground finch. R2 . R2-1_GStR2 non-LTR retrotransposon Gavia stellata from red-throated loon. R2 .R2-1_HAl R2 non-LTR retrotransposon Haliaeetus from white-tailed eagle.albicilla R2 . R2-1_IS R2 non-LTR retrotransposon Ixodes scapularis fromdeer tick. R2 . R2-1_LCh R2-type non-LTR Latimeria retrotransposon -consensus. chalumnae R2 . R2-1_LDi R2 non-LTR retrotransposon Leptosomusfrom cuckoo roller. discolor R2 . R2-1_LSal non-LTR retrotransposon,Lepeophtheirus consensus. salmonis R2 . R2-1_LV R2 non-LTRretrotransposon Lytechinus from green sea urchin. variegatus R2 .R2-1_MDe R2 non-LTR retrotransposon Mayetiola from Hessian fly.destructor R2 . R2-1_MLe R2 non-LTR retrotransposon - Mnemiopsis leidyiconsensus. R2 . R2-1_MR R2 non-LTR retrotransposon Megachile fromalfalfa leafcutter bee. rotundata R2 . R2-1_MUn R2 non-LTRretrotransposon Melopsittacus fragment from budgerigar. undulatus R2 .R2-1_MUni R2 non-LTR retrotransposon Mesitornis from brown mesite.unicolor R2 . R2-1_MVi R2 non-LTR retrotransposon Manacus vitellinusfrom golden-collared manakin. R2 . R2-1_NNi R2 non-LTR retrotransposonNipponia nippon from created ibis. R2 . R2-1_NV Starlet sea anemoneR2-1_NV Nematostella autonomous Non-LTR vectensis Retrotransposon -consensus. R2 . R2-1_OHo R2 non-LTR retrotransposon Opisthocomus fromhoatzin. hoazin R2 . R2-1_PAd R2 non-LTR retrotransposon Pygoscelisadeliae from Adelie penguin. R2 . R2-1_PBa R2 non-LTR retrotransposonPogonomyrmex from red harvester ant. barbatus R2 . R2-1_PCar R2 non-LTRretrotransposon Phalacrocorax from great cormorant. carbo R2 . R2-1_PCauR2 non-LTR retrotransposon Priapulus caudatus sequence. R2 . R2-1_PCr R2non-LTR retrotransposon Podiceps cristatus from great crested grebe. R2. R2-1_PCri R2 non-LTR retrotransposon Pelecanus crispus from Dalmatianpelican. R2 . R2-1_PGu R2 non-LTR retrotransposon Pterocles fromsandgrouse. gutturalis R2 . R2-1_PLe R2 non-LTR retrotransposon Phaethonlepturus from tropicbird. R2 . R2-1_PM R2-1_PM is a family of R2Petromyzon marinus non-LTR retrotransposons - consensus. R2 . R2-1_PPapR2 non-LTR retrotransposon Phlebotomus from sand fly. papatasi R2 .R2-1_PPu R2 non-LTR retrotransposon Picoides pubescens from downywoodpecker. R2 . R2-1_PRR R2 non-LTR retrotransposon Phoenicopterus fromAmerican flamingo. ruber ruber R2 . R2-1_PSi R2 non-LTR retrotransposonPelodiscus from Chinese soft-shelled sinensis turtle. R2 . R2-1_RMi R2non-LTR retrotransposon Rhipicephaus from brown tick. microplus R2 .R2-1_RPr R2 non-LTR retrotransposon Rhodnius prolixus sequence. R2 .R2-1_RPu R2 non-LTR retrotransposon Rhipicephalus cDNA sequence frombrown tick. pulchellus R2 . R2-1_SCa R2 non-LTR retrotransposon Serinuscanaria from Atlantic canary. R2 . R2-1_SK R2 non-LTR retrotransposonSaccoglossus from acorn worm. kowalevskii R2 . R2-1_SM R2-typeretrotransposon from Schmidtea Schmidtea mediterranea: mediterraneaconsensus. R2 . R2-1_SP R2 non-LTR retrotransposon Strongylocentrotusfrom purple sea urchin. purpuratus R2 AGKD01072455 R2-1_SSa R2-typenon-LTR retrotransposon. Salmo salar R2 . R2-1_StC R2 non-LTRretrotransposon Struthiocamelus from ostrich. australis R2 . R2-1_TAl R2non-LTR retrotransposon Tyto alba from barn owl. R2 . R2-1_TCas R2non-LTR retrotransposon Tribolium from red flour beetle - consensuscastaneum R2 . R2-1_TG A family of R2 non-LTR Taeniopygiaretrotransposons - consensus guttata sequence. R2 . R2-1_TGut R2 non-LTRretrotransposon Tinamus guttatus from white-throated tinamou. R2 .R2-1_TSP A family of R2 non-LTR Trichinella retrotransposons in thespiralis Trichinella spiralis genome - a consensus. R2 scaffold_6R2-1_TUr R2 non-LTR retrotransposon Tetranychus from twospotted spidermite. urticae R2 . R2-1_XM R2 non-LTR retrotransposon Xiphophorusfragment from Southern maculatus platyfish. R2 . R2-1_ZA R2 non-LTRretrotransposon Zonotrichia from white-throated sparrow. albicollis R2 .R2-1_ZLM R2 non-LTR retrotransposon Zosterops from silvereye. lateralismelanops R2 . R2-2_APi R2 non-LTR retrotransposon Acyrthosiphon from peaaphid. pisum R2 . R2-2_CCan R2 non-LTR retrotransposon Cuculus canorusfrom common cuckoo. R2 . R2-2_CMa R2 non-LTR retrotransposon Chlamydotisfrom Macqueen's bustard. macqueenii R2 . R2-2_DWi 28S rDNA-specificnon-LTR Drosophila retrotransposon R2 in willistoni Drosophilawillistoni. R2 . R2-2_HAl R2 non-LTR retrotransposon Haliaeetus fromwhite-tailed eagle. albicilla R2 . R2-2_IS R2 non-LTR retrotransposonIxodes scapularis from deer tick. R2 . R2-2_MR R2 non-LTRretrotransposon Megachile from alfalfa leafcutter bee. rotundata R2 .R2-2_MUn R2 non-LTR retrotransposon Melopsittacus fragment frombudgerigar. undulatus R2 . R2-2_MUni R2 non-LTR retrotransposonMesitornis from brown mesite. unicolor R2 . R2-2_NNi R2 non-LTRretrotransposon Nipponia nippon from created ibis. R2 . R2-2_NV Starletsea anemone R2-2_NV Nematostella autonomous Non-LTR vectensisRetrotransposon - consensus. R2 . R2-2_PBa R2 non-LTR retrotransposonPogonomyrmex from red harvester ant. barbatus R2 . R2-2_PM R2-2_PM is afamily of R2 Petromyzon marinus non-LTR retrotransposons - a consensus.R2 . R2-2_RPr R2 non-LTR retrotransposon Rhodnius prolixus sequence. R2. R2-2_SMed R2 non-LTR retrotransposon Schmidtea from Schmidteamediterranea: mediterranea consensus. R2 . R2-2_TCas R2 non-LTRretrotransposon Tribolium from red flour beetle. castaneum R2scaffold_37 R2-2_TUr R2 non-LTR retrotransposon Tetranychus fromtwospotted spider mite. urticae R2 ABJB010555169 R2-3_IS R2 non-LTRretrotransposon Ixodes scapularis from deer tick. R2 . R2-3_MR R2non-LTR retrotransposon Megachile from alfalfa leafcutter bee. rotundataR2 . R2-4_MR R2 non-LTR retrotransposon Megachile from alfalfaleafcutter bee. rotundata R2 . R2-5_MR R2 non-LTR retrotransposonMegachile from alfalfa leafcutter bee. rotundata R2 . R2-6_MR R2 non-LTRretrotransposon Megachile from alfalfa leafcutter bee. rotundata R2 .R2-7_MR R2 non-LTR retrotransposon Megachile from alfalfa leafcutterbee. rotundata R2 . R2-8_MR R2 non-LTR retrotransposon Megachile fromalfalfa leafcutter bee. rotundata R2 . R2-N1_Gav Non-LTRretrotransposon. Gavialis gangeticus R2 . R2-N2_Gav Non-LTRretrotransposon. Gavialis gangeticus R2 . R2-N2B_Gav Non-LTRretrotransposon. Gavialis gangeticus R2 . R2A_NVi 28S rDNA-specificnon-LTR Nasonia retrotransposon R2 in vitripennis Nasonia vitripennis.R2 AF015817 R2A_TM Tenebrio molitor Tenebrio molitor retrotransposon R2reverse transcriptase gene, partial cds. R2 . R2Amel R2Amel - R2 non-LTRApis mellifera retrotransposon from the honeybee Apis mellifera. R2AF015685 R2B_DM Drosophila mercatorum R2 Drosophila retrotransposonreverse mercatorum transcriptase domain protein gene, complete cds. R2 .R2B_NVi 28S rDNA-specific non-LTR Nasonia retrotransposon R2 invitripennis Nasonia vitripennis. R2 AF015822 R2B_TM Tenebrio molitorTenebrio molitor retrotransposon R2 reverse transcriptase gene, partialcds. R2 . R2C_NGi 28S rDNA-specific non-LTR Nasonia giraultiretrotransposon R2 in Nasonia giraulti. R2 AB097122 R2Ci-B Cionaintestinalis Ciona intestinalis retrotransposon R2Ci-B, completesequence. R2 . R2Ci-D Ciona intestinalis Ciona intestinalisretrotransposon R2CiD, complete sequence. R2 AB097121 R2CIA_CI Cionaintestinalis Ciona intestinalis retrotransposon R2Ci-A, completesequence. R2 AB097125 R2Cs-D Ciona intestinalis Ciona savignyiretrotransposon R2CsD, partial sequence. R2 . R2D_NGi 28S rDNA-specificnon-LTR Nasonia giraulti retrotransposon R2 in Nasonia giraulti. R2NM_001030097 R2Dr R2 non-LTR retrotransposon in Danio rerio the Daniorerio genome - a single copy. R2 . R2E_NLo 28S rDNA-specific non-LTRNasonia retrotransposon R2 in longicornis Nasonia longicornisi. R2AB201408 R2Eb R2 non-LTR retrotransposon Eptatretus burgeri fromEptatretus burgeri. R2 AB201415 R2Ha R2 non-LTR retrotransposon Hasariusadansoni from Hasarius adansoni. R2 JN937617 R2La R2-type non-LTRretrotransposon. Lepidurus arcticus R2 . R2LcA R2-type non-LTRretrotransposon. Lepidurus couesii R2 JN937619 R2LcB R2-type non-LTRretrotransposon. Lepidurus couesii R2 . R2LcC R2-type non-LTRretrotransposon. Lepidurus couesii R2 JN937616 R2Ll R2-type non-LTRretrotransposon. Lepidurus apus lubbocki R2 AB201414 R2Mr R2 non-LTRretrotransposon Metacrinus from Metacrinus rotundus. rotundus R2 .R2NS-1_CGi R2-type retrotransposon from Crassostrea gigas Crassostreagigas. R2 . R2NS-1_CSi R2-type retrotransposon from ClonorchisClonorchis sinensis: consensus. sinensis R2 . R2NS-1_PMi R2-like non-LTRPatiria miniata retrotransposon from bat star. R2 . R2NS-1_SMed R2-typeretrotransposon from Schmidtea Schmidtea mediterranea: mediterraneaconsensus. R2 . R2Nvec-A R2Nvec-A - R2 non-LTR Nematostellaretrotransposon from the vectensis starlet sea anemone Nematostellavectensis. R2 . R2Ol-A R2 non-LTR retrotransposon Oryzias latipes fromthe medaka Oryzias latipes - consensus. R2 AB201416 R2Pc R2 non-LTRretrotransposon Procambarus from Procambarus clarkii. clarkii R2 .R2Sm-A R2Sm-A - R2 non-LTR Schistosoma retrotransposon from the mansonibloodfluke Schistosoma mansoni. R2 AB201409 R2Ta R2 non-LTRretrotransposon Tanichthys from Tanichthys albonubes. albonubes R2EU854578 R2Tc R2-type non-LTR retrotransposon. Triops cancriformis R2JN937621 R2Tc_it R2-type non-LTR retrotransposon. Triops cancriformis R2AB201417 R2Tl R2 non-LTR retrotransposon Triops from Triopslongicaudatus. longicaudatus R4 U29445 R4_AL Ascaris lumbricoidesAscaris site-specific non-LTR lumbricoides retrotransposable element R4in 26S rDNA, complete sequence. R4 U29590 R4_HC Haemonchus contortusnon-LTR Haemonchus retrotransposon specific to contortus the largesubunit rRNA genes of nematodes. R4 . R4_Hmel a R4 element fromHeliconius Heliconius melpomene. melpomene R4 . R4-1_AC A family of R4non-LTR Anolis retrotransposons - consensus carolinensis sequence. R4 .R4-1_ADi R4-type retrotransposon: Acropora consensus. digitifera R4 .R4-1_BM Non-LTR retrotransposon - a Bombyx mori consensus. R4CADV01008175 R4-1_BX An R4 non-LTR retrotransposon Bursaphelenchusfamily from Bursaphelenchus xylophilus xylophilus. R4 ABLE03011482R4-1_CJap An R4 non-LTR retrotransposon Caenorhabditis family fromCaenorhabditis japonica japonica. R4 . R4-1_CM Non-LTR retrotransposonfrom Callorhinchus the elephant shark - consensus. milii R4 . R4-1_CPBNon-LTR retrotransposon: Chrysemyspicta consensus. bellii R4 . R4-1_EDAutonomous non-LTR Entamoeba dispar retrotransposon from the R4 clade -a consensus sequence. R4 . R4-1_HG An R4 non-LTR retrotransposonHeterodera family from glycines Heterodera glycines. R4 . R4-1_HMeNon-LTR retrotransposon family from Heliconius Heliconius melpomenemelpomene. melpomene melpomene R4 CABB01003843 R4-1_MI An R4 non-LTRretrotransposon Meloidogyne family from Meloidogyne incognita incognita.R4 . R4-1_PH Non-LTR Retrotransposon, Parhyale consensus. hawaiensis R4CACX01002001 R4-1_SRa An R4 non-LTR retrotransposon Strongyloides familyfrom Strongyloides ratti. ratti R4 . R4-1_TCa R4-type retrotransposon:Tribolium consensus. castaneum R4 . R4-1B_AC Dong-type non-LTR Anolisretrotransposons - a consensus carolinensis sequence. R4 . R4-2_AS An R4non-LTR retrotransposon Ascaris suum family from Ascaris suum. R4CADV01009048 R4-2_BX An R4 non-LTR retrotransposon Bursaphelenchusfamily from Bursaphelenchus xylophilus xylophilus. R4 ABLA01000389R4-2_HG An R4 non-LTR retrotransposon Heterodera family from Heteroderaglycines glycines. R4 CACX01002006 R4-2_SRa An R4 non-LTRretrotransposon Strongyloides family from Strongyloides ratti. ratti R4CADV01008832 R4-3_BX An R4 non-LTR retrotransposon Bursaphelenchusfamily from Bursaphelenchus xylophilus xylophilus. R4 . R4-3_SRa An R4non-LTR retrotransposon Strongyloides family from Strongyloides ratti.ratti R4 . R4-4_BX An R4 non-LTR retrotransposon Bursaphelenchus familyfrom Bursaphelenchus xylophilus xylophilus. R4 . R4-4_SRa An R4 non-LTRretrotransposon Strongyloides family from Strongyloides ratti ratti. R4. R4-5_BX An R4 non-LTR retrotransposon Bursaphelenchus family fromBursaphelenchus xylophilus xylophilus. NeSL AY216701 R5 Girardia tigrinaR5 Girardia tigrina retrotransposon, complete sequence. NeSL . R5-1_SM Afamily of planarian NeSL Schmidtea non-LTR retrotransposons -mediterranea consensus. NeSL . R5-2_SM A family of planarian NeSLSchmidtea non-LTR retrotransposons - mediterranea consensus. R2 . R8Hm-AR8Hm-A - 18S rDNA-specific Hydra vulgaris non-LTR retrotransposon fromHydra magnipapillata. R2 . R8Hm-B R8Hm-B - 18S rDNA-specific Hydravulgaris non-LTR retrotransposon from Hydra magnipapillata. R2 . R9AvR9Av, an rDNA-specific non-LTR Adineta vaga retrotransposon family fromrotifer. R2 FJ461304 RaR2 28S rDNA-specific non-LTR Rhynchosciararetrotransposon R2 from americana Rhynchosciara americana. R4 . Rex6Non-LTR retrotransposon; Takifugu rubripes site-specific LINE; R4/Dongsuperfamily; REX6; DONG_FR. R4 . Rex6-1_OL A Rex6 non-LTRretrotransposon Oryzias latipes family from Olyzias latipes. CRE X17078SLACS Trypanosoma brucei DNA for Trypanosoma brucei retrotransposableelement SLACS. NeSL . Utopia-1_ACa Utopia-1_ACa is a protozoanAcanthamoeba Utopia non-LTR retrotransposon - castellanii a completesequence. NeSL scaffold_474 Utopia-1_ACar A family of NeSL non-LTRAnolis retrotransposons. carolinensis NeSL . Utopia-1_AEc A family ofUtopia non-LTR Acromyrmex retrotransposons - consensus. echinatior NeSL. Utopia-1_AMi A family of NeSL non-LTR Alligator retrotransposons -consensus. mississippiensis NeSL . Utopia-1_APi A family of Utopianon-LTR Acyrthosiphon retrotransposons - consensus. pisum NeSL .Utopia-1_APl A family of Utopia non-LTR Agrilus retrotransposons.planipennis NeSL . Utopia-1_CFl A family of Utopia non-LTR Camponotusretrotransposons - consensus. floridanus NeSL . Utopia-1_CMy A family ofUtopia non-LTR Chelonia mydas retrotransposons - consensus. NeSL .Utopia-1_CPB A family of Utopia non-LTR Chrysemyspictaretrotransposons - consensus. bellii NeSL . Utopia-1_Crp Non-LTRretrotransposon. Crocodylus porosus NeSL . Utopia-1_DPo A family ofUtopia non-LTR Dendroctonus ponderosae retrotransposons. NeSL .Utopia-1_DPu A family of Utopia non-LTR Daphnia pulex retrotransposons -consensus. NeSL . Utopia-1_DYak A family of Utopia non-LTR Drosophilayakuba retrotransposons - consensus. NeSL . Utopia-1_EBr A family ofUtopia non-LTR Eimeria brunetti retrotransposons - consensus. NeSL .Utopia-1_EMi A family of Utopia non-LTR Eimeria mitis retrotransposons -consensus. NeSL . Utopia-1_ENe A family of Utopia non-LTR Eimerianecatrix retrotransposons - consensus. NeSL . Utopia-1_Gav Non-LTRretrotransposon. Gavialis gangeticus NeSL . Utopia-1_GG1 A family ofUtopia non-LTR Ganaspis retrotransposons. NeSL . Utopia-1_HAra A familyof Utopia non-LTR Hyaloperonospora retrotransposons. arabidopsidis NeSL. Utopia-1_HG A family of Utopia non-LTR Heterodera retrotransposons.glycines NeSL . Utopia-1_HMM A family of Utopia non-LTR Heliconiusretrotransposons. melpomene melpomene NeSL . Utopia-1_HSal A family ofUtopia non-LTR Harpegnathos retrotransposons - consensus. saltator NeSL. Utopia-1_IS A family of Utopia non-LTR Ixodes scapularisretrotransposons - consensus. NeSL . Utopia-1_LAl A family of Utopianon-LTR Lasioglossum retrotransposons. albipes NeSL . Utopia-1_LFu Afamily of Utopia non-LTR Ladona fulva retrotransposons. NeSLAGCV01358106 Utopia-1_LV A family of Utopia non-LTR Lytechinusretrotransposons. variegatus NeSL . Utopia-1_MRo A family of Utopianon-LTR Megachile retrotransposons - consensus. rotundata NeSL .Utopia-1_NVit A family of Utopia non-LTR Nasonia retrotransposons -consensus. vitripennis NeSL . Utopia-1_PAlni NeSL non-LTRretrotransposon Phytophthora alni from Phytophthora alni. NeSL .Utopia-1_PArrh NeSL non-LTR retrotransposon Pythium from Pythiumarrhenomanes. arrhenomanes NeSL . Utopia-1_PBa A family of Utopianon-LTR Pogonomyrmex retrotransposons - consensus. barbatus NeSL .Utopia-1_PCa A family of Utopia non-LTR Phytophthora retrotransposons -consensus. capsici NeSL . Utopia-1_PCinn NeSL non-LTR retrotransposonPhytophthora from Phytophthora cinnamomi. cinnamomi NeSL AHJF01004292Utopia-1_PCu A family of Utopia non-LTR Pseudoperonosporaretrotransposons - consensus. cubensis NeSL . Utopia-1_PI A family ofNeSL non-LTR Phytophthora retrotransposons - consensus. infestans NeSL .Utopia-1_PInsi NeSL non-LTR retrotransposon Pythium insidiosum fromPythium insidiosum. NeSL . Utopia-1_PKern NeSL non-LTR retrotransposonPhytophthora from Phytophthora kernoviae. kernoviae NeSL .Utopia-1_PLate NeSL non-LTR retrotransposon Phytophthora fromPhytophthora lateralis. lateralis NeSL . Utopia-1_PMi A family of Utopianon-LTR Patiria miniata retrotransposons. NeSL . Utopia-1_PPac A familyof Utopia non-LTR Pristionchus retrotransposons. pacificus NeSL .Utopia-1_PPini NeSL non-LTR retrotransposon Phytophthora from pinifoliaPhytophthora pinifolia. NeSL . Utopia-1_PR A family of Utopia non-LTRPhytophthora retrotransposons - consensus. ramorum NeSL . Utopia-1_PRe Afamily of Utopia non-LTR Panagrellus retrotransposons - consensus.redivivus NeSL . Utopia-1_PS A family of Utopia non-LTR Phytophthorasojae retrotransposons - consensus. NeSL . Utopia-1_PSi A family ofUtopia non-LTR Pelodiscus retrotransposons - consensus. sinensis NeSL .Utopia-1_PT A family of Utopia non-LTR Parasteatoda retrotransposons.tepidariorum NeSL ADOS01001321 Utopia-1_PU A family of Utopia non-LTRPythium ultimum retrotransposons. NeSL . Utopia-1_PVexa NeSL non-LTRretrotransposon Phytopythium from Phytopythium vexans. aff. vexans NeSL. Utopia-1_SaPa A family of Utopia non-LTR Saprolegnia retrotransposons.parasitica NeSL . Utopia-1_SDicl NeSL non-LTR retrotransposonSaprolegnia from Saprolegnia diclina. diclina NeSL . Utopia-1_SM Afamily of Utopia non-LTR Strigamia maritima retrotransposons. NeSLAAGJ02140537 Utopia-1_SP A family of Utopia non-LTR Strongylocentrotusretrotransposons. purpuratus NeSL . Utopia-1_TSP A family of Utopianon-LTR Trichinella retrotransposons. spiralis NeSL . Utopia-1B_CPB Afamily of Utopia non-LTR Chrysemys picta retrotransposons - consensus.bellii NeSL . Utopia-2_APi A family of Utopia non-LTR Acyrthosiphonretrotransposons. pisum NeSL . Utopia-2_CMy A family of Utopia non-LTRChelonia mydas retrotransposons - consensus. NeSL . Utopia-2_CPB Afamily of Utopia non-LTR Chrysemys picta retrotransposons - consensus.bellii NeSL . Utopia-2_DPu A family of Utopia non-LTR Daphnia pulexretrotransposons. NeSL . Utopia-2_LFu A family of Utopia non-LTR Ladonafulva retrotransposons. NeSL . Utopia-2_PCa A family of Utopia non-LTRPhytophthora retrotransposons - consensus. capsici NeSL . Utopia-2_PI Afamily of NeSL non-LTR Phytophthora retrotransposons - consensus.infestans NeSL . Utopia-2_PR A family of Utopia non-LTR Phytophthoraretrotransposons - consensus. ramorum NeSL . Utopia-2_PS A family ofUtopia non-LTR Phytophthora sojae retrotransposons - consensus. NeSL .Utopia-2_PU A family of Utopia non-LTR Pythium ultimum retrotransposons.NeSL . Utopia-3_CPB A family of Utopia non-LTR Chrysemys pictaretrotransposons - consensus. bellii NeSL . Utopia-3_DPu A family ofUtopia non-LTR Daphnia pulex retrotransposons. NeSL . Utopia-3_LFu Afamily of Utopia non-LTR Ladona fulva retrotransposons. NeSL .Utopia-3_PCa A family of Utopia non-LTR Phytophthora retrotransposons -consensus. capsici NeSL . Utopia-3_PI A family of NeSL non-LTRPhytophthora retrotransposons - consensus. infestans NeSL . Utopia-3_PRA family of Utopia non-LTR Phytophthora retrotransposons - consensus.ramorum NeSL . Utopia-4_LFu A family of Utopia non-LTR Ladona fulvaretrotransposons. NeSL AATU01001281.1 Utopia-4_PI A family of NeSLnon-LTR Phytophthora retrotransposons - a copy. infestans NeSL .Utopia-4_PR A family of Utopia non-LTR Phytophthora retrotransposons -consensus. ramorum NeSL . Utopia-5_LFu A family of Utopia non-LTR Ladonafulva retrotransposons. NeSL . Utopia-5_PI A family of Utopia non-LTRPhytophthora retrotransposons - consensus. infestans NeSL . Utopia-5_PRA family of Utopia non-LTR Phytophthora retrotransposons - consensus.ramorum NeSL . Utopia-6_LFu A family of Utopia non-LTR Ladona fulvaretrotransposons. R4 . X4_LINE Conserved LINE element Vertebratareconstructed from the human genome - consensus. NeSL . YURE_CSa A NeSLnon-LTR retrotransposon Ciona savignyi from Ciona savignyi. R2 .YURE-2_Cis YURE non-LTR retrotransposon Ciona savignyi from Cionasavignyi. NeSL . YURECi Ciona intestinalis Ciona intestinalisretrotransposon YURECi.

A skilled artisan can, based on the Accession numbers provided in Tables1-3 determine the nucleic acid and corresponding polypeptide sequencesof each retrotransposon and domains thereof, e.g., by using routinesequence analysis tools as Basic Local Alignment Search Tool (BLAST) orCD-Search for conserved domain analysis. Other sequence analysis toolsare known and can be found, e.g., at https://molbiol-tools.ca, forexample, at https://molbiol-tools.ca/Motifs.htm. SEQ ID NOs 1-112 alignwith each row in Table 1, and SEQ ID NOs 113-1015 align with the first903 rows of Table 2.

Tables 1-3 herein provide the sequences of exemplary transposons,including the amino acid sequence of the retrotransposase, and sequencesof 5′ and 3′ untranslated regions to allow the retrotransposase to bindthe template RNA, and the full transposon nucleic acid sequence. In someembodiments, a 5′ UTR of any of Tables 1-3 allows the retrotransposaseto bind the template RNA. In some embodiments, a 3′ UTR of any of Tables1-3 allows the retrotransposase to bind the template RNA. Thus, in someembodiments, a polypeptide for use in any of the systems describedherein can be a polypeptide of any of Tables 1-3 herein, or a sequencehaving at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%identity thereto. In some embodiments, the system further comprises oneor both of a 5′ or 3′ untranslated region of any of Tables 1-3 herein(or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%,98%, or 99% identity thereto), e.g., from the same transposon as thepolypeptide referred to in the preceding sentence, as indicated in thesame row of the same table. In some embodiments, the system comprisesone or both of a 5′ or 3′ untranslated region of any of Tables 1-3herein, e.g., a segment of the full transposon sequence that encodes anRNA that is capable of binding a retrotransposase, and/or thesub-sequence provided in the column entitled Predicted 5′ UTR orPredicted 3′ UTR.

In some embodiments, a polypeptide for use in any of the systemsdescribed herein can be a molecular reconstruction or ancestralreconstruction based upon the aligned polypeptide sequence of multipleretrotransposons. In some embodiments, a 5′ or 3′ untranslated regionfor use in any of the systems described herein can be a molecularreconstruction based upon the aligned 5′ or 3′ untranslated region ofmultiple retrotransposons. A skilled artisan can, based on the Accessionnumbers provided herein, align polypeptides or nucleic acid sequences,e.g., by using routine sequence analysis tools as Basic Local AlignmentSearch Tool (BLAST) or CD-Search for conserved domain analysis.Molecular reconstructions can be created based upon sequence consensus,e.g. using approaches described in Ivies et al., Cell 1997, 501-510;Wagstaff et al., Molecular Biology and Evolution 2013, 88-99. In someembodiments, the retrotransposon from which the 5′ or 3′ untranslatedregion or polypeptide is derived is a young or a recently active mobileelement, as assessed via phylogenetic methods such as those described inBoissinot et al., Molecular Biology and Evolution 2000, 915-928.

Table 3 (below) shows exemplary Gene Writer proteins and associatedsequences from a variety of retrotransposases, identified using datamining. Column 1 indicates the family to which the retrotransposonbelongs. Column 2 lists the element name. Column 3 indicates anaccession number, if any. Column 4 lists an organism in which theretrotransposase is found. Column 5 lists the DNA sequence of theretrotransposon. Column 6 lists the predicted 5′ untranslated region,and column 7 lists the predicted 3′ untranslated region; both aresegments of the sequence of column 5 that are predicted to allow thetemplate RNA to bind the retrotransposase of column 8. (It is understoodthat columns 5-7 show the DNA sequence, and that an RNA sequenceaccording to any of columns 5-7 would typically include uracil ratherthan thymidine.) Column 8 lists the predicted retrotransposase sequenceencoded in the retrotransposon of column 5.

Lengthy table referenced here US20200109398A1-20200409-T00001 Pleaserefer to the end of the specification for access instructions.

Gene Writers, e.g. Thermostable Gene Writers

While not wishing to be bound by theory, in some embodiments,retrotransposases that evolved in cold environments may not function aswell at human body temperature. This application provides a number ofthermostable Gene Writers, including proteins derived from avianretrotransposases. Exemplary avian transposase sequences in Table 3include those of Taeniopygia guttata (zebra finch; transposon nameR2-1_TG), Geospiza fortis (medium ground finch; transposon nameR2-1_Gfo), Zonotrichia albicollis (white-throated sparrow; transposonname R2-1_ZA), and Tinamus guttatus (white-throated tinamou; transposonname R2-1_TGut).

Thermostability may be measured, e.g., by testing the ability of a GeneWriter to polymerize DNA in vitro at a high temperature (e.g., 37° C.)and a low temperature (e.g., 25° C.). Suitable conditions for assayingin vitro DNA polymerization activity (e.g., processivity) are described,e.g., in Bibillo and Eickbush, “High Processivity of the ReverseTranscriptase from a Non-long Terminal Repeat Retrotransposon” (2002)JBC 277, 34836-34845. In some embodiments, the thermostable Gene Writerpolypeptide has an activity, e.g., a DNA polymerization activity, at 37°C. that is no less than 70%, 75%, 80%, 85%, 90%, or 95% of its activityat 25° C. under otherwise similar conditions.

In some embodiments, a GeneWriter polypeptide (e.g., a sequence of Table1, 2, or 3 or a sequence having at least 70%, 75%, 80%, 85%, 90%, 95%,96%, 97%, 98%, or 99% identity thereto) is stable in a subject chosenfrom a mammal (e.g., human) or a bird. In some embodiments, a GeneWriterpolypeptide described herein is functional at 37° C. In someembodiments, a GeneWriter polypeptide described herein has greateractivity at 37° C. than it does at a lower temperature, e.g., at 30° C.,25° C., or 20° C. In some embodiments, a GeneWriter polypeptidedescribed herein has greater activity in a human cell than in azebrafish cell.

In some embodiments, a GeneWriter polypeptide is active in a human cellcultured at 37° C., e.g., using an assay of Example 6 or Example 7herein.

In some embodiments, the assay comprises steps of: (1) introducingHEK293T cells into one or more wells of 6.4 mm diameter, at 10,000cells/well, (2) incubating the cells at 37° C. for 24 hr, (3) providinga transfection mixture comprising 0.5 μl if FuGENE® HD transfectionreagent and 80 ng DNA (wherein the DNA is a plasmid comprising, inorder, (a) CMV promoter, (b) 100 bp of sequence homologous to the 100 bpupstream of the target site, (c) sequence encoding a 5′ untranslatedregion that binds the GeneWriter protein, (d) sequence encoding theGeneWriter protein, (e) sequence encoding a 3′ untranslated region thatbinds the GeneWriter protein (f) 100 bp of sequence homologous to the100 bp downstream of the target site, and (g) BGH polyadenylationsequence) and 10 μl Opti-MEM and incubating for 15 min at roomtemperature, (4) adding the transfection mixture to the cells, (5)incubating the cells for 3 days, and (6) assaying integration of theexogenous sequence into a target locus (e.g., rDNA) in the cell genome,e.g., wherein one or more of the preceding steps are performed asdescribed in Example 6 herein.

In some embodiments, the GeneWriter polypeptide results in insertion ofthe heterologous object sequence (e.g., the GFP gene) into the targetlocus (e.g., rDNA) at an average copy number of at least 0.01, 0.025,0.05, 0.075, 0.1, 0.15, 0.2, 0.25, 0.3, 0.4, 0.5, 0.75, 1, 1.25, 1.5,1.75, 2, 2.5, 3, 4, or 5 copies per genome. In some embodiments, a celldescribed herein (e.g., a cell comprising a heterologous sequence at atarget insertion site) comprises the heterologous object sequence at anaverage copy number of at least 0.01, 0.025, 0.05, 0.075, 0.1, 0.15,0.2, 0.25, 0.3, 0.4, 0.5, 0.75, 1, 1.25, 1.5, 1.75, 2, 2.5, 3, 4, or 5copies per genome.

In some embodiments, a GeneWriter causes integration of a sequence in atarget RNA with relatively few truncation events at the terminus. Forinstance, in some embodiments, a Gene Writer protein (e.g., of SEQ IDNO: 1016) results in about 25-100%, 50-100%, 60-100%, 70-100%, 75-95%,80%-90%, or 86.17% of integrants into the target site beingnon-truncated, as measured by an assay described herein, e.g., an assayof Example 6 and FIG. 8. In some embodiments, a Gene Writer protein(e.g., of SEQ ID NO: 1016) results in at least about 30%, 40%, 50%, 60%,70%, 80%, or 90% of integrants into the target site being non-truncated,as measured by an assay described herein. In some embodiments, anintegrant is classified as truncated versus non-truncated using an assaycomprising amplification with a forward primer situated 565 bp from theend of the element (e.g., a wild-type transposon sequence, e.g., ofTaeniopygia guttata) and a reverse primer situated in the genomic DNA ofthe target insertion site, e.g., rDNA. In some embodiments, the numberof full-length integrants in the target insertion site is greater thanthe number of integrants truncated by 300-565 nucleotides in the targetinsertion site, e.g., the number of full-length integrants is at least1.1×, 1.2×, 1.5×, 2×, 3×, 4×, 5×, 6×, 7×, 8×, 9×, or 10× the number ofthe truncated integrants, or the number of full-length integrants is atleast 1.1×-10×, 2×-10×, 3×-10×, or 5×-10× the number of the truncatedintegrants.

In some embodiments, a system or method described herein results ininsertion of the heterologous object sequence only at one target site inthe genome of the target cell. Insertion can be measured, e.g., using athreshold of above 1%, 1.5%, 2%, 2.5%, 3%, 3.5%, 4%, 4.5%, 5%, e.g., asdescribed in Example 8. In some embodiments, a system or methoddescribed herein results in insertion of the heterologous objectsequence wherein less than 1%, 1.5%, 2%, 2.5%, 3%, 3.5%, 4%, 4.5%, 5%,10%, 20%, 30%, 40%, or 50% of insertions are at a site other than thetarget site, e.g., using an assay described herein, e.g., an assay ofExample 8.

In some embodiments, a system or method described herein results in“scarless” insertion of the heterologous object sequence, while in someembodiments, the target site can show deletions or duplications ofendogenous DNA as a result of insertion of the heterologous sequence.The mechanisms of different retrotransposons could result in differentpatterns of duplications or deletions in the host genome occurringduring retrotransposition at the target site. In some embodiments, thesystem results in a scarless insertion, with no duplications ordeletions in the surrounding genomic DNA. In some embodiments, thesystem results in a deletion of less than 1, 2, 3, 4, 5, 10, 50, or 100bp of genomic DNA upstream of the insertion. In some embodiments, thesystem results in a deletion of less than 1, 2, 3, 4, 5, 10, 50, or 100bp of genomic DNA downstream of the insertion. In some embodiments, thesystem results in a duplication of less than 1, 2, 3, 4, 5, 10, 50, or100 bp of genomic DNA upstream of the insertion. In some embodiments,the system results in a duplication of less than 1, 2, 3, 4, 5, 10, 50,or 100 bp of genomic DNA downstream of the insertion.

In some embodiments, a GeneWriter described herein, or a DNA-bindingdomain thereof, binds to its target site specifically, e.g., as measuredusing an assay of Example 21. In some embodiments, the GeneWriter orDNA-binding domain thereof binds to its target site more strongly thanto any other binding site in the human genome. For example, in someembodiments, in an assay of Example 21, the target site represents morethan 50%, 60%, 70%, 80%, 90%, or 95% of binding events of the GeneWriteror DNA-binding domain thereof to human genomic DNA.

Genetically Engineered, e.g., Dimerized GeneWriters

Some non-LTR retrotransposons utilize two subunits to completeretrotransposition (Christensen et al PNAS 2006). In some embodiments, aretrotransposase described herein comprises two connected subunits as asingle polypeptide. For instance, two wild-type retrotransposases couldbe joined with a linker to form a covalently “dimerized” protein (seeFIG. 17). In some embodiments, the nucleic acid coding for theretrotransposase codes for two retrotransposase subunits to be expressedas a single polypeptide. In some embodiments, the subunits are connectedby a peptide linker, such as has been described herein in the sectionentitled “Linker” and, e.g., in Chen et al Adv Drug Deliv Rev 2013. Insome embodiments, the two subunits in the polypeptide are connected by arigid linker. In some embodiments, the rigid linker consists of themotif (EAAAK)_(n) (SEQ ID NO: 1534). In other embodiments, the twosubunits in the polypeptide are connected by a flexible linker. In someembodiments, the flexible linker consists of the motif (Gly)_(n). Insome embodiments, the flexible linker consists of the motif (GGGGS)_(n)(SEQ ID NO: 1535). In some embodiments, the rigid or flexible linkerconsists of 1, 2, 3, 4, 5, 10, 15, or more amino acids in length toenable retrotransposition. In some embodiments, the linker consists of acombination of rigid and flexible linker motifs.

Based on mechanism, not all functions are required from bothretrotransposase subunits. In some embodiments, the fusion protein mayconsist of a fully functional subunit and a second subunit lacking oneor more functional domains. In some embodiments, one subunit may lackreverse transcriptase functionality. In some embodiments, one subunitmay lack the reverse transcriptase domain. In some embodiments, onesubunit may possess only endonuclease activity. In some embodiments, onesubunit may possess only an endonuclease domain. In some embodiments,the two subunits comprising the single polypeptide may providecomplimentary functions.

In some embodiments, one subunit may lack endonuclease functionality. Insome embodiments, one subunit may lack the endonuclease domain. In someembodiments, one subunit may possess only reverse transcriptaseactivity. In some embodiments, one subunit may possess only a reversetranscriptase domain. In some embodiments, one subunit may possess onlyDNA-dependent DNA synthesis functionality.

Linkers:

In some embodiments, domains of the compositions and systems describedherein (e.g., the endonuclease and reverse transcriptase domains of apolypeptide or the DNA binding domain and reverse transcriptase domainsof a polypeptide) may be joined by a linker. A composition describedherein comprising a linker element has the general form 1-L-S2, whereinS1 and S2 may be the same or different and represent two domain moieties(e.g., each a polypeptide or nucleic acid domain) associated with oneanother by the linker. In some embodiments, a linker may connect twopolypeptides. In some embodiments, a linker may connect two nucleic acidmolecules. In some embodiments, a linker may connect a polypeptide and anucleic acid molecule. A linker may be a chemical bond, e.g., one ormore covalent bonds or non-covalent bonds. A linker may be flexible,rigid, and/or cleavable. In some embodiments, the linker is a peptidelinker. Generally, a peptide linker is at least 2, 3, 4, 5, 6, 7, 8, 9,10 or more amino acids in length, e.g., 2-50 amino acids in length, 2-30amino acids in length.

The most commonly used flexible linkers have sequences consistingprimarily of stretches of Gly and Ser residues (“GS” linker). Flexiblelinkers may be useful for joining domains that require a certain degreeof movement or interaction and may include small, non-polar (e.g. Gly)or polar (e.g. Ser or Thr) amino acids. Incorporation of Ser or Thr canalso maintain the stability of the linker in aqueous solutions byforming hydrogen bonds with the water molecules, and therefore reduceunfavorable interactions between the linker and the other moieties.Examples of such linkers include those having the structure [GGS]^(≥1)or [GGGS]^(≥1) (SEQ ID NO: 1536). Rigid linkers are useful to keep afixed distance between domains and to maintain their independentfunctions. Rigid linkers may also be useful when a spatial separation ofthe domains is critical to preserve the stability or bioactivity of oneor more components in the agent. Rigid linkers may have an alphahelix-structure or Pro-rich sequence, (XP)n, with X designating anyamino acid, preferably Ala, Lys, or Glu. Cleavable linkers may releasefree functional domains in vivo. In some embodiments, linkers may becleaved under specific conditions, such as the presence of reducingreagents or proteases. In vivo cleavable linkers may utilize thereversible nature of a disulfide bond. One example includes athrombin-sensitive sequence (e.g., PRS) between the two Cys residues. Invitro thrombin treatment of CPRSC (SEQ ID NO: 1537) results in thecleavage of the thrombin-sensitive sequence, while the reversibledisulfide linkage remains intact. Such linkers are known and described,e.g., in Chen et al. 2013. Fusion Protein Linkers: Property, Design andFunctionality. Adv Drug Deliv Rev. 65(10): 1357-1369. In vivo cleavageof linkers in compositions described herein may also be carried out byproteases that are expressed in vivo under pathological conditions (e.g.cancer or inflammation), in specific cells or tissues, or constrainedwithin certain cellular compartments. The specificity of many proteasesoffers slower cleavage of the linker in constrained compartments.

In some embodiments the amino acid linkers are (or are homologous to)the endogenous amino acids that exist between such domains in a nativepolypeptide. In some embodiments the endogenous amino acids that existbetween such domains are substituted but the length is unchanged fromthe natural length. In some embodiments, additional amino acid residuesare added to the naturally existing amino acid residues between domains.

In some embodiments, the amino acid linkers are designed computationallyor screened to maximize protein function (Anad et al., FEBS Letters,587:19, 2013).

Template RNA Component of Gene Writer™ Gene Editor System

The Gene Writer systems described herein can transcribe an RNA sequencetemplate into host target DNA sites by target-primed reversetranscription. By writing DNA sequence(s) via reverse transcription ofthe RNA sequence template directly into the host genome, the Gene Writersystem can insert an object sequence into a target genome without theneed for exogenous DNA sequences to be introduced into the host cell(unlike, for example, CRISPR systems), as well as eliminate an exogenousDNA insertion step. Therefore, the Gene Writer system provides aplatform for the use of customized RNA sequence templates containingobject sequences, e.g., sequences comprising heterologous gene codingand/or function information.

In some embodiments the template RNA encodes a Gene Writer protein incis with a heterologous object sequence. Various cis constructs weredescribed, for example, in Kuroki-Kami et al (2019) Mobile DNA 10:23(incorporated by reference herein in its entirety), and can be used incombination with any of the embodiments described herein. For instance,in some embodiments, the template RNA comprises a heterologous objectsequence, a sequence encoding a Gene Writer protein (e.g., a proteincomprising (i) a reverse transcriptase domain and (ii) an endonucleasedomain, e.g., as described herein), a 5′ untranslated region, and a 3′untranslated region. The components may be included in various orders.In some embodiments, the Gene Writer protein and heterologous objectsequence are encoded in different directions (sense vs. anti-sense),e.g., using an arrangement shown in FIG. 3A of Kuroki-Kami et al, Id. Insome embodiments the Gene Writer protein and heterologous objectsequence are encoded in the same direction. In some embodiments, thenucleic acid encoding the polypeptide and the template RNA or thenucleic acid encoding the template RNA are covalently linked, e.g., arepart of a fusion nucleic acid and/or are part of the same transcript. Insome embodiments, the fusion nucleic acid comprises RNA or DNA.

The nucleic acid encoding the Gene Writer polypeptide may, in someinstances, be 5′ of the heterologous object sequence. For example, insome embodiments, the template RNA comprises, from 5′ to 3′, a 5′untranslated region, a sense-encoded Gene Writer polypeptide, asense-encoded heterologous object sequence, and 3′ untranslated region.In some embodiments, the template RNA comprises, from 5′ to 3′, a 5′untranslated region, a sense-encoded Gene Writer polypeptide,anti-sense-encoded heterologous object sequence, and 3′ untranslatedregion.

In some embodiments, the RNA further comprises homology to the DNAtarget site.

It is understood that, when a template RNA is described as comprising anopen reading frame or the reverse complement thereof, in someembodiments the template RNA must be converted into double stranded DNA(e.g., through reverse transcription) before the open reading frame canbe transcribed and translated.

In certain embodiments, customized RNA sequence template can beidentified, designed, engineered and constructed to contain sequencesaltering or specifying host genome function, for example by introducinga heterologous coding region into a genome; affecting or causing exonstructure/alternative splicing; causing disruption of an endogenousgene; causing transcriptional activation of an endogenous gene; causingepigenetic regulation of an endogenous DNA; causing up- ordown-regulation of operably liked genes, etc. In certain embodiments, acustomized RNA sequence template can be engineered to contain sequencescoding for exons and/or transgenes, provide for binding sites totranscription factor activators, repressors, enhancers, etc., andcombinations of thereof. In other embodiments, the coding sequence canbe further customized with splice acceptor sites, poly-A tails. Incertain embodiments the RNA sequence can contain sequences coding for anRNA sequence template homologous to the RLE transposase, be engineeredto contain heterologous coding sequences, or combinations thereof.

The template RNA may have some homology to the target DNA. In someembodiments the template RNA has at least 2, 3, 4, 5, 6, 7, 8, 9, 10,11, 12, 13, 14, 15, 20, 25, 30, 35, 40, 45, 50, 60, 70, 80, 90, 100,110, 120, 130, 140, 150, 175, 200 or more bases of exact homology to thetarget DNA at the 3′ end of the RNA. In some embodiments the templateRNA has at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 20, 25,30, 35, 40, 45, 50, 60, 70, 80, 90, 100, 110, 120, 130, 140, 150, 160,175, 180, or 200 or more bases of at least 50%, 60%, 70%, 80%, 85%, 90%,95%, 97%, 98%, 99% or 100% homology to the target DNA, e.g., at the 5′end of the template RNA. In some embodiments the template RNA has a 3′untranslated region derived from a non-LTR retrotransposon, e.g. anon-LTR retrotransposons described herein. In some embodiments thetemplate RNA has a 3′ region of at least 10, 15, 20, 25, 30, 40, 50, 60,80, 100, 120, 140, 160, 180, 200 or more bases of at least 50%, 60%,70%, 80%, 85%, 90%, 95%, 97%, 98%, 99% or 100% homology to the 3′sequence of a non-LTR retrotransposon, e.g., a non-LTR retrotransposondescribed herein, e.g. a non-LTR retrotransposon in Table 1, 2, or 3. Insome embodiments the template RNA has a 5′ untranslated region derivedfrom a non-LTR retrotransposon, e.g. a non-LTR retrotransposonsdescribed herein. In some embodiments the template RNA has a 5′ regionof at least 10, 15, 20, 25, 30, 40, 50, 60, 80, 100, 120, 140, 160, 180,or 200 or more bases of at least 40%, 50%, 60%, 70%, 80%, 90%, 95% orgreater homology to the 5′ sequence of a non-LTR retrotransposon, e.g.,a non-LTR retrotransposon described herein, e.g. a non-LTRretrotransposon described in Table 2 or 3.

The template RNA component of a Gene Writer genome editing systemdescribed herein typically is able to bind the Gene Writer genomeediting protein of the system. In some embodiments the template RNA hasa 3′ region that is capable of binding a Gene Writer genome editingprotein. The binding region, e.g., 3′ region, may be a structured RNAregion, e.g., having at least 1, 2 or 3 hairpin loops, capable ofbinding the Gene Writer genome editing protein of the system.

The template RNA component of a Gene Writer genome editing systemdescribed herein typically is able to bind the Gene Writer genomeediting protein of the system. In some embodiments the template RNA hasa 5′ region that is capable of binding a Gene Writer genome editingprotein. The binding region, e.g., 5′ region, may be a structured RNAregion, e.g., having at least 1, 2 or 3 hairpin loops, capable ofbinding the Gene Writer genome editing protein of the system. In someembodiments, the 5′ untranslated region comprises a pseudoknot, e.g., apseudoknot that is capable of binding to the Gene Writer protein.

In some embodiments, the template RNA (e.g., an untranslated region ofthe hairpin RNA, e.g., a 5′ untranslated region) comprises a stem-loopsequence. In some embodiments, the template RNA (e.g., an untranslatedregion of the hairpin RNA, e.g., a 5′ untranslated region) comprises ahairpin. In some embodiments, the template RNA (e.g., an untranslatedregion of the hairpin RNA, e.g., a 5′ untranslated region) comprises ahelix. In some embodiments, the template RNA (e.g., an untranslatedregion of the hairpin RNA, e.g., a 5′ untranslated region) comprises apsuedoknot. In some embodiments the template RNA comprises a ribozyme.In some embodiments the ribozyme is similar to an hepatitis delta virus(HDV) ribozyme, e.g., has a secondary structure like that of the HDVribozyme and/or has one or more activities of the HDV ribozyme, e.g., aself-cleavage activity. See, e.g., Eickbush et al., Molecular andCellular Biology, 2010, 3142-3150.

In some embodiments, the template RNA (e.g., an untranslated region ofthe hairpin RNA, e.g., a 3′ untranslated region) comprises one or morestem-loops or helices. Exemplary structures of R2 3′ UTRs are shown, forexample, in Ruschak et al. “Secondary structure models of the 3′untranslated regions of diverse R2 RNAs” RNA. 2004 June; 10(6): 978-987,e.g., at FIG. 3, therein, and in Eikbush and Eikbush, “R2 and R2/R1hybrid non-autonomous retrotransposons derived by internal deletions offull-length elements” Mobile DNA (2012) 3:10; e.g., at FIG. 3 therein,which articles are hereby incorporated by reference in their entirety.

In some embodiments, a template RNA described herein comprises asequence that is capable of binding to a GeneWriter protein describedherein. For instance, in some embodiments, the template RNA comprises anMS2 RNA sequence capable of binding to an MS2 coat protein sequence inthe GeneWriter protein. In some embodiments, the template RNA comprisesan RNA sequence capable of binding to a B-box sequence. In someembodiments, the template RNA comprises an RNA sequence (e.g., a crRNAsequence and/or tracrRNA sequence) capable of binding to a dCas sequencein the GeneWriter protein. In some embodiments, in addition to or inplace of a UTR, the template RNA is linked (e.g., covalently) to anon-RNA UTR, e.g., a protein or small molecule.

In some embodiments the template RNA has a poly-A tail at the 3′ end. Insome embodiments the template RNA does not have a poly-A tail at the 3′end.

In some embodiments the template RNA has a 5′ region of at least 10, 15,20, 25, 30, 40, 50, 60, 80, 100, 120, 140, 160, 180, 200 or more basesof at least 40%, 50%, 60%, 70%, 80%, 90%, 95% or greater homology to the5′ sequence of a non-LTR retrotransposon, e.g., a non-LTRretrotransposon described herein.

The template RNA of the system typically comprises an object sequencefor insertion into a target DNA. The object sequence may be coding ornon-coding.

In some embodiments a system or method described herein comprises asingle template RNA. In some embodiments a system or method describedherein comprises a plurality of template RNAs.

In some embodiments, the object sequence may contain an open readingframe. In some embodiments the template RNA has a Kozak sequence. Insome embodiments the template RNA has an internal ribosome entry site.In some embodiments the template RNA has a self-cleaving peptide such asa T2A or P2A site. In some embodiments the template RNA has a startcodon. In some embodiments the template RNA has a splice acceptor site.In some embodiments the template RNA has a splice donor site. In someembodiments the template RNA has a microRNA binding site downstream ofthe stop codon. In some embodiments the template RNA has a polyA taildownstream of the stop codon of an open reading frame. In someembodiments the template RNA comprises one or more exons. In someembodiments the template RNA comprises one or more introns. In someembodiments the template RNA comprises a eukaryotic transcriptionalterminator. In some embodiments the template RNA comprises an enhancedtranslation element or a translation enhancing element. In someembodiments the RNA comprises the human T-cell leukemia virus (HTLV-1) Rregion. In some embodiments the RNA comprises a posttranscriptionalregulatory element that enhances nuclear export, such as that ofHepatitis B Virus (HPRE) or Woodchuck Hepatitis Virus (WPRE). In someembodiments, in the template RNA, the heterologous object sequenceencodes a polypeptide and is coded in an antisense direction withrespect to the 5′ and 3′ UTR. In some embodiments, in the template RNA,the heterologous object sequence encodes a polypeptide and is coded in asense direction with respect to the 5′ and 3′ UTR.

In some embodiments, a nucleic acid described herein (e.g., a templateRNA or a DNA encoding a template RNA) comprises a microRNA binding site.In some embodiments, the microRNA binding site is used to increase thetarget-cell specificity of a GeneWriter system. For instance, themicroRNA binding site can be chosen on the basis that is is recognizedby a miRNA that is present in a non-target cell type, but that is notpresent (or is present at a reduced level relative to the non-targetcell) in a target cell type. Thus, when the template RNA is present in anon-target cell, it would be bound by the miRNA, and when the templateRNA is present in a target cell, it would not be bound by the miRNA (orbound but at reduced levels relative to the non-target cell). While notwishing to be bound by theory, binding of the miRNA to the template RNAmay interfere with insertion of the heterologous object sequence intothe genome. Accordingly, the heterologous object sequence would beinserted into the genome of target cells more efficiently than into thegenome of non-target cells. A system having a microRNA binding site inthe template RNA (or DNA encoding it) may also be used in combinationwith a nucleic acid encoding a GeneWriter polypeptide, whereinexpression of the GeneWriter polypeptide is regulated by a secondmicroRNA binding site, e.g., as described herein, e.g., in the sectionentitled “Polypeptide component of Gene Writer gene editor system”.

In some embodiments, the object sequence may contain a non-codingsequence. For example, the template RNA may comprise a promoter orenhancer sequence. In some embodiments the template RNA comprises atissue specific promoter or enhancer, each of which may beunidirectional or bidirectional. In some embodiments the promoter is anRNA polymerase I promoter, RNA polymerase II promoter, or RNA polymeraseIII promoter. In some embodiments the promoter comprises a TATA element.In some embodiments the promoter comprises a B recognition element. Insome embodiments the promoter has one or more binding sites fortranscription factors. In some embodiments the non-coding sequence istranscribed in an antisense-direction with respect to the 5′ and 3′ UTR.In some the non-coding sequence is transcribed in a sense direction withrespect to the 5′ and 3′ UTR.

In some embodiments, a nucleic acid described herein (e.g., a templateRNA or a DNA encoding a template RNA) comprises a promoter sequence,e.g., a tissue specific promoter sequence. In some embodiments, thetissue-specific promoter is used to increase the target-cell specificityof a GeneWriter system. For instance, the promoter can be chosen on thebasis that it is active in a target cell type but not active in (oractive at a lower level in) a non-target cell type. Thus, even if thepromoter integrated into the genome of a non-target cell, it would notdrive expression (or only drive low level expression) of an integratedgene. A system having a tissue-specific promoter sequence in thetemplate RNA may also be used in combination with a microRNA bindingsite, e.g., in the template RNA or a nucleic acid encoding a GeneWriterprotein, e.g., as described herein. A system having a tissue-specificpromoter sequence in the template RNA may also be used in combinationwith a DNA encoding a GeneWriter polypeptide, driven by atissue-specific promoter, e.g., to achieve higher levels of GeneWriterprotein in target cells than in non-target cells.

In some embodiments the template RNA comprises a microRNA sequence, asiRNA sequence, a guide RNA sequence, a piwi RNA sequence.

In some embodiments the template RNA comprises a site that coordinatesepigenetic modification. In some embodiments the template RNA comprisesan element that inhibits, e.g., prevents, epigenetic silencing. In someembodiments the template RNA comprises a chromatin insulator. Forexample, the template RNA comprises a CTCF site or a site targeted forDNA methylation.

In order to promote higher level or more stable gene expression, thetemplate RNA may include features that prevent or inhibit genesilencing. In some embodiments, these features prevent or inhibit DNAmethylation. In some embodiments, these features promote DNAdemethylation. In some embodiments, these features prevent or inhibithistone deacetylation. In some embodiments, these features prevent orinhibit histone methylation. In some embodiments, these features promotehistone acetylation. In some embodiments, these features promote histonedemethylation. In some embodiments, multiple features may beincorporated into the template RNA to promote one or more of thesemodifications. CpG dinculeotides are subject to methylation by hostmethyl transferases. In some embodiments, the template RNA is depletedof CpG dinucleotides, e.g., does not comprise CpG nucleotides orcomprises a reduced number of CpG dinucleotides compared to acorresponding unaltered sequence. In some embodiments, the promoterdriving transgene expression from integrated DNA is depleted of CpGdinucleotides.

In some embodiments the template RNA comprises a gene expression unitcomposed of at least one regulatory region operably linked to aneffector sequence. The effector sequence may be a sequence that istranscribed into RNA (e.g., a coding sequence or a non-coding sequencesuch as a sequence encoding a micro RNA).

In some embodiments the object sequence of the template RNA is insertedinto a target genome in an endogenous intron. In some embodiments theobject sequence of the template RNA is inserted into a target genome andthereby acts as a new exon. In some embodiments the insertion of theobject sequence into the target genome results in replacement of anatural exon or the skipping of a natural exon.

In some embodiments the object sequence of the template RNA is insertedinto the target genome in a genomic safe harbor site, such as AAVS1,CCR5, or ROSA26. In some embodiment the object sequence of the templateRNA is added to the genome in an intergenic or intragenic region. Insome embodiments the object sequence of the template RNA is added to thegenome 5′ or 3′ within 0.1 kb, 0.25 kb, 0.5 kb, 0.75, kb, 1 kb, 2 kb, 3kb, 4 kb, 5 kb, 7.5 kb, 10 kb, 15 kb, 20 kb, 25 kb, 50, 75 kb, or 100 kbof an endogenous active gene. In some embodiments the object sequence ofthe template RNA is added to the genome 5′ or 3′ within 0.1 kb, 0.25 kb,0.5 kb, 0.75, kb, 1 kb, 2 kb, 3 kb, 4 kb, 5 kb, 7.5 kb, 10 kb, 15 kb, 20kb, 25 kb, 50, 75 kb, or 100 kb of an endogenous promoter or enhancer.In some embodiments the object sequence of the template RNA can be,e.g., 50-50,000 base pairs (e.g., between 50-40,000 bp, between500-30,000 bp between 500-20,000 bp, between 100-15,000 bp, between500-10,000 bp, between 50-10,000 bp, between 50-5,000 bp. In someembodiments, the heterologous object sequence is less than 1,000, 1,300,1500, 2,000, 3,000, 4,000, 5,000, or 7,500 nucleotides in length.

In some embodiments the genomic safe harbor site is a Natural Harbor™site. In some embodiments the Natural Harbor™ site is ribosomal DNA(rDNA). In some embodiments the Natural Harbor™ site is 5S rDNA, 18SrDNA, 5.8S rDNA, or 28S rDNA. In some embodiments the Natural Harbor™site is the Mutsu site in 5S rDNA. In some embodiments the NaturalHarbor™ site is the R2 site, the R5 site, the R6 site, the R4 site, theR1 site, the R9 site, or the RT site in 28S rDNA. In some embodimentsthe Natural Harbor™ site is the R8 site or the R7 site in 18S rDNA. Insome embodiments the Natural Harbor™ site is DNA encoding transfer RNA(tRNA). In some embodiments the Natural Harbor™ site is DNA encodingtRNA-Asp or tRNA-Glu. In some embodiments the Natural Harbor™ site isDNA encoding spliceosomal RNA. In some embodiments the Natural Harbor™site is DNA encoding small nuclear RNA (snRNA) such as U2 snRNA.

Thus, in some aspects, the present disclosure provides a method ofinserting a heterologous object sequence into a Natural Harbor™ site. Insome embodiments, the method comprises using a GeneWriter systemdescribed herein, e.g., using a polypeptide of any of Tables 1-3 or apolypeptide having sequence similarity thereto, e.g., at least 80%, 85%,90%, or 95% identity thereto. In some embodiments, the method comprisesusing an enzyme, e.g., a retrotransposase, to insert the heterologousobject sequence into the Natural Harbor™ site. In some aspects, thepresent disclosure provides a host human cell comprising a heterologousobject sequence (e.g., a sequence encoding a therapeutic polypeptide)situated at a Natural Harbor™ site in the genome of the cell. In someembodiments, the Natural Harbor™ site is a site described in Table 4below. In some embodiments, the heterologous object sequence is insertedwithin 20, 50, 100, 150, 200, 250, 500, or 1000 base pairs of a sequenceshown in Table 4. In some embodiments, the heterologous object sequenceis inserted within 0.1 kb, 0.25 kb, 0.5 kb, 0.75, kb, 1 kb, 2 kb, 3 kb,4 kb, 5 kb, 7.5 kb, 10 kb, 15 kb, 20 kb, 25 kb, 50, 75 kb, or 100 kb ofa sequence shown in Table 4. In some embodiments, the heterologousobject sequence is inserted into a site having at least 70%, 75%, 80%,85%, 90%, 95%, 96%, 97%, 98%, or 99% identity to a sequence shown inTable 4. In some embodiments, the heterologous object sequence isinserted within 20, 50, 100, 150, 200, 250, 500, or 1000 base pairs, orwithin 0.1 kb, 0.25 kb, 0.5 kb, 0.75, kb, 1 kb, 2 kb, 3 kb, 4 kb, 5 kb,7.5 kb, 10 kb, 15 kb, 20 kb, 25 kb, 50, 75 kb, or 100 kb, of a sitehaving at least 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%identity to a sequence shown in Table 4. In some embodiments, theheterologous object sequence is inserted within a gene indicated inColumn 5 of Table 4, or within 20, 50, 100, 150, 200, 250, 500, or 1000base pairs, or within 0.1 kb, 0.25 kb, 0.5 kb, 0.75, kb, 1 kb, 2 kb, 3kb, 4 kb, 5 kb, 7.5 kb, 10 kb, 15 kb, 20 kb, 25 kb, 50, 75 kb, or 100kb, of the gene.

TABLE 4 Natural Harbor ™ sites. Column 1 indicates a retrotransposonthat inserts into the Natural Harbor ™ site. Column 2indicates the gene at the Natural Harbor ™ site. Columns 3and 4 show exemplary human genome sequence 5′ and 3′ of theinsertion site (for example, 250 bp). Columns 5 and 6 listthe example gene symbol and corresponding Gene ID. Example Target TargetGene Example Site Gene 5′ flanking sequence 3′ flanking sequence SymbolGene ID R2 28S rDNA CCGGTCCCCCCCGCCGGGTCC GTAGCCAAATGCCTCGTCATC RNA28SN1106632264 GCCCCCGGGGCCGCGGTTCC TAATTAGTGACGCGCATGAATGCGCGGCGCCTCGCCTCGGC GGATGAACGAGATTCCCACT CGGCGCCTAGCAGCCGACTTGTCCCTACCTACTATCCAGCG AGAACTGGTGCGGACCAGGG AAACCACAGCCAAGGGAACGGAATCCGACTGTTTAATTAAA GGCTTGGCGGAATCAGCGGG ACAAAGCATCGCGAAGGCCCGAAAGAAGACCCTGTTGAGC GCGGCGGGTGTTGACGCGAT TTGACTCTAGTCTGGCACGGTGTGATTTCTGCCCAGTGCTCT GAAGAGACATGAGAGGTGTA GAATGTCAAAGTGAAGAAATGAATAAGTGGGAGGCCCCCG TCAATGAAGCGCGGGTAAAC GCGCCCCCCCGGTGTCCCCGCGGCGGGAGTAACTATGACTC GAGGGGCCCGGGGCGGGGT TCTTAAG (SEQ ID NO: 1508)CCGCCG (SEQ ID NO: 1513) R4 28S rDNA GCGGTTCCGCGCGGCGCCTCCGCATGAATGGATGAACGAG RNA28SN1 106632264 GCCTCGGCCGGCGCCTAGCAATTCCCACTGTCCCTACCTACT GCCGACTTAGAACTGGTGCG ATCCAGCGAAACCACAGCCAGACCAGGGGAATCCGACTGT AGGGAACGGGCTTGGCGGA TTAATTAAAACAAAGCATCGCATCAGCGGGGAAAGAAGACC GAAGGCCCGCGGCGGGTGTT CTGTTGAGCTTGACTCTAGTCGACGCGATGTGATTTCTGCCC TGGCACGGTGAAGAGACATG AGTGCTCTGAATGTCAAAGTAGAGGTGTAGAATAAGTGGG GAAGAAATTCAATGAAGCGC AGGCCCCCGGCGCCCCCCCGGGGTAAACGGCGGGAGTAAC GTGTCCCCGCGAGGGGCCCG TATGACTCTCTTAAGGTAGCCGGGCGGGGTCCGCCGGCCCT AAATGCCTCGTCATCTAATTA GCGGGCCGCCGGTGAAATACGTGACG (SEQ ID NO: 1509) CACTACTC (SEQ ID NO: 1514) R5 28S rDNATCCCCCCCGCCGGGTCCGCCC CCAAATGCCTCGTCATCTAAT RNA28SN1 106632264CCGGGGCCGCGGTTCCGCGC TAGTGACGCGCATGAATGGA GGCGCCTCGCCTCGGCCGGCTGAACGAGATTCCCACTGTCC GCCTAGCAGCCGACTTAGAA CTACCTACTATCCAGCGAAACCTGGTGCGGACCAGGGGAAT CACAGCCAAGGGAACGGGCT CCGACTGTTTAATTAAAACAATGGCGGAATCAGCGGGGAAA AGCATCGCGAAGGCCCGCGG GAAGACCCTGTTGAGCTTGACGGGTGTTGACGCGATGTGA CTCTAGTCTGGCACGGTGAA TTTCTGCCCAGTGCTCTGAATGAGACATGAGAGGTGTAGAA GTCAAAGTGAAGAAATTCAA TAAGTGGGAGGCCCCCGGCGTGAAGCGCGGGTAAACGGCG CCCCCCCGGTGTCCCCGCGAG GGAGTAACTATGACTCTCTTAGGGCCCGGGGCGGGGTCCG AGGTAG (SEQ ID NO: 1510) CCGGCCC (SEQ ID NO: 1515)R9 28S rDNA CGGCGCGCTCGCCGGCCGAG TAGCTGGTTCCCTCCGAAGTT RNA28SN1106632264 GTGGGATCCCGAGGCCTCTC TCCCTCAGGATAGCTGGCGCTCAGTCCGCCGAGGGCGCACC CTCGCAGACCCGACGCACCCC ACCGGCCCGTCTCGCCCGCCGCGCCACGCAGTTTTATCCGGT CGCCGGGGAGGTGGAGCAC AAAGCGAATGATTAGAGGTCGAGCGCACGTGTTAGGACCC TTGGGGCCGAAACGATCTCA GAAAGATGGTGAACTATGCCACCTATTCTCAAACTTTAAAT TGGGCAGGGCGAAGCCAGA GGGTAAGAAGCCCGGCTCGCGGAAACTCTGGTGGAGGTCC TGGCGTGGAGCCGGGCGTGG GTAGCGGTCCTGACGTGCAAAATGCGAGTGCCTAGTGGGC ATCGGTCGTCCGACCTGGGT CACTTTTGGTAAGCAGAACTGATAGGGGCGAAAGACTAATC GCGCTGCGGGATGAACCGAA GAACCATCTAG (SEQ ID NO:CGCC (SEQ ID NO: 1516) 1511) R8 18S rDNA GCATTCGTATTGCGCCGCTAGTGAAACTTAAAGGAATTGAC RNA18SN1 106631781 AGGTGAAATTCTTGGACCGGGGAAGGGCACCACCAGGAGT CGCAAGACGGACCAGAGCGA GGAGCCTGCGGCTTAATTTGAAGCATTTGCCAAGAATGTTT ACTCAACACGGGAAACCTCA TCATTAATCAAGAACGAAAGTCCCGGCCCGGACACGGACAG CGGAGGTTCGAAGACGATCA GATTGACAGATTGATAGCTCTGATACCGTCGTAGTTCCGACC TTCTCGATTCCGTGGGTGGTG ATAAACGATGCCGACCGGCGGTGCATGGCCGTTCTTAGTTG ATGCGGCGGCGTTATTCCCAT GTGGAGCGATTTGTCTGGTTGACCCGCCGGGCAGCTTCCG AATTCCGATAACGAACGAGA GGAAACCAAAGTCTTTGGGTCTCTGGCATGCTAACTAGTTA TCCGGGGGGAGTATGGTTGC CGCGACCCCCGAGCGGTCGGAAAGC (SEQ ID NO: 1512) CGTCCC (SEQ ID NO: 1517) R4-2_SRa tRNA-AspTRD-GTC1-1 100189207 LIN25_SM tRNA-Glu TRE-CTC1-1 100189384 R1 28S rDNATAGCAGCCGACTTAGAACTG ACCTACTATCCAGCGAAACCA RNA28SN1 106632264GTGCGGACCAGGGGAATCCG CAGCCAAGGGAACGGGCTTG ACTGTTTAATTAAAACAAAGCGCGGAATCAGCGGGGAAAG ATCGCGAAGGCCCGCGGCGG AAGACCCTGTTGAGCTTGACTGTGTTGACGCGATGTGATTTC CTAGTCTGGCACGGTGAAGA TGCCCAGTGCTCTGAATGTCAGACATGAGAGGTGTAGAATA AAGTGAAGAAATTCAATGAA AGTGGGAGGCCCCCGGCGCCGCGCGGGTAAACGGCGGGA CCCCCGGTGTCCCCGCGAGG GTAACTATGACTCTCTTAAGGGGCCCGGGGCGGGGTCCGCC TAGCCAAATGCCTCGTCATCT GGCCCTGCGGGCCGCCGGTGAATTAGTGACGCGCATGAAT AAATACCACTACTCTGATCGT GGATGAACGAGATTCCCACTTTTTTCACTGACCCGGTGAGG GTCCCT (SEQ ID NO: 1518) CGGGGGG (SEQ ID NO: 1524)R6 28S rDNA CCCCCCGCCGGGTCCGCCCCC AAATGCCTCGTCATCTAATTA RNA28SN1106632264 GGGGCCGCGGTTCCGCGCGG GTGACGCGCATGAATGGATG CGCCTCGCCTCGGCCGGCGCAACGAGATTCCCACTGTCCCT CTAGCAGCCGACTTAGAACT ACCTACTATCCAGCGAAACCAGGTGCGGACCAGGGGAATCC CAGCCAAGGGAACGGGCTTG GACTGTTTAATTAAAACAAAGGCGGAATCAGCGGGGAAAG CATCGCGAAGGCCCGCGGCG AAGACCCTGTTGAGCTTGACTGGTGTTGACGCGATGTGATT CTAGTCTGGCACGGTGAAGA TCTGCCCAGTGCTCTGAATGTGACATGAGAGGTGTAGAATA CAAAGTGAAGAAATTCAATG AGTGGGAGGCCCCCGGCGCCAAGCGCGGGTAAACGGCGG CCCCCGGTGTCCCCGCGAGG GAGTAACTATGACTCTCTTAAGGCCCGGGGCGGGGTCCGCC GGTAGCC (SEQ ID NO: 1519) GGCCCTG (SEQ ID NO: 1525)R7 18S rDNA GCGCAAGACGGACCAGAGCG GGAGCCTGCGGCTTAATTTG RNA18SN1 106631781AAAGCATTTGCCAAGAATGTT ACTCAACACGGGAAACCTCA TTCATTAATCAAGAACGAAAGCCCGGCCCGGACACGGACAG TCGGAGGTTCGAAGACGATC GATTGACAGATTGATAGCTCTAGATACCGTCGTAGTTCCGAC TTCTCGATTCCGTGGGTGGTG CATAAACGATGCCGACCGGCGTGCATGGCCGTTCTTAGTTG GATGCGGCGGCGTTATTCCC GTGGAGCGATTTGTCTGGTTATGACCCGCCGGGCAGCTTC AATTCCGATAACGAACGAGA CGGGAAACCAAAGTCTTTGGCTCTGGCATGCTAACTAGTTA GTTCCGGGGGGAGTATGGTT CGCGACCCCCGAGCGGTCGGGCAAAGCTGAAACTTAAAGG CGTCCCCCAACTTCTTAGAGG AATTGACGGAAGGGCACCACGACAAGTGGCGTTCAGCCAC CAGGAGT (SEQ ID NO: 1520) CCGAG (SEQ ID NO: 1526)RT 28S rDNA GGCCGGGCGCGACCCGCTCC AACTGGCTTGTGGCGGCCAA RNA28SN1 106632264GGGGACAGTGCCAGGTGGG GCGTTCATAGCGACGTCGCTT GAGTTTGACTGGGGCGGTACTTTGATCCTTCGATGTCGGCT ACCTGTCAAACGGTAACGCA CTTCCTATCATTGTGAAGCAGGGTGTCCTAAGGCGAGCTCA AATTCACCAAGCGTTGGATTG GGGAGGACAGAAACCTCCCGTTCACCCACTAATAGGGAACG TGGAGCAGAAGGGCAAAAG TGAGCTGGGTTTAGACCGTCCTCGCTTGATCTTGATTTTCA GTGAGACAGGTTAGTTTTACC GTACGAATACAGACCGTGAACTACTGATGATGTGTTGTTGC AGCGGGGCCTCACGATCCTTC CATGGTAATCCTGCTCAGTACTGACCTTTTGGGTTTTAAGCA GAGAGGAACCGCAGGTTCAG GGAGGTGTCAGAAAAGTTACACATTTGGTGTATGTGCTTGG CACAGGGAT (SEQ ID NO: C (SEQ ID NO: 1527) 1521)Mutsu 5S rDNA GTCTACGGCCATACCACCC TGAACGCGCCCGATCTCGTCT RNA5S1 100169751(SEQ ID NO: 1522) GATCTCGGAAGCTAAGCAGG GTCGGGCCTGGTTAGTACTTGGATGGGAGACCGCCTGGGA ATACCGGGTGCTGTAGGCTTT (SEQ ID NO: 1528) Utopia/U2 snRNA ATCGCTTCTCGGCCTTTTGGC TCTGTTCTTATCAGTTTAATAT RNU2-1      6066Keno TAAGATCAAGTGTAGTA (SEQ CTGATACGTCCTCTATCCGAG ID NO: 1523)GACAATATATTAAATGGATTT TTGGAGCAGGGAGATGGAAT AGGAGCTTGCTCCGTCCACTCCACGCATCGACCTGGTATTGC AGTACCTCCAGGAACGGTGC ACCC (SEQ ID NO: 1529)

In some embodiments, a system or method described herein results ininsertion of a heterologous sequence into a target site in the humangenome. In some embodiments, the target site in the human genome hassequence similarity to the corresponding target site of thecorresponding wild-type retrotransposase (e.g., the retrotransposasefrom which the GeneWriter was derived) in the genome of the organism towhich it is native. For instance, in some embodiments, the identitybetween the 40 nucleotides of human genome sequence centered at theinsertion site and the 40 nucleotides of native organism genome sequencecentered at the insertion site is less than 99.5%, 99%, 98%, 97%, 96%,95%, 90%, 85%, 80%, 75%, 70%, 60%, or 50%, or is between 50-60%, 60-70%,70-80%, 80-90%, or 90-100%. In some embodiments, the identity betweenthe 100 nucleotides of human genome sequence centered at the insertionsite and the 100 nucleotides of native organism genome sequence centeredat the insertion site is less than 99.5%, 99%, 98%, 97%, 96%, 95%, 90%,85%, 80%, 75%, 70%, 60%, or 50%, or is between 50-60%, 60-70%, 70-80%,80-90%, or 90-100%. In some embodiments, the identity between the 500nucleotides of human genome sequence centered at the insertion site andthe 500 nucleotides of native organism genome sequence centered at theinsertion site is less than 99.5%, 99%, 98%, 97%, 96%, 95%, 90%, 85%,80%, 75%, 70%, 60%, or 50%, or is between 50-60%, 60-70%, 70-80%,80-90%, or 90-100%.

Production of Compositions and Systems

As will be appreciated by one of skill, methods of designing andconstructing nucleic acid constructs and proteins or polypeptides (suchas the systems, constructs and polypeptides described herein) areroutine in the art. Generally, recombinant methods may be used. See, ingeneral, Smales & James (Eds.), Therapeutic Proteins: Methods andProtocols (Methods in Molecular Biology), Humana Press (2005); andCrommelin, Sindelar & Meibohm (Eds.), Pharmaceutical Biotechnology:Fundamentals and Applications, Springer (2013). Methods of designing,preparing, evaluating, purifying and manipulating nucleic acidcompositions are described in Green and Sambrook (Eds.), MolecularCloning: A Laboratory Manual (Fourth Edition), Cold Spring HarborLaboratory Press (2012).

Exemplary methods for producing a therapeutic pharmaceutical protein orpolypeptide described herein involve expression in mammalian cells,although recombinant proteins can also be produced using insect cells,yeast, bacteria, or other cells under control of appropriate promoters.Mammalian expression vectors may comprise non-transcribed elements suchas an origin of replication, a suitable promoter, and other 5′ or 3′flanking non-transcribed sequences, and 5′ or 3′ non-translatedsequences such as necessary ribosome binding sites, a polyadenylationsite, splice donor and acceptor sites, and termination sequences. DNAsequences derived from the SV40 viral genome, for example, SV40 origin,early promoter, splice, and polyadenylation sites may be used to provideother genetic elements required for expression of a heterologous DNAsequence. Appropriate cloning and expression vectors for use withbacterial, fungal, yeast, and mammalian cellular hosts are described inGreen & Sambrook, Molecular Cloning: A Laboratory Manual (FourthEdition), Cold Spring Harbor Laboratory Press (2012).

Various mammalian cell culture systems can be employed to express andmanufacture recombinant protein. Examples of mammalian expressionsystems include CHO, COS, HEK293, HeLA, and BHK cell lines. Processes ofhost cell culture for production of protein therapeutics are describedin Zhou and Kantardjieff (Eds.), Mammalian Cell Cultures for BiologicsManufacturing (Advances in Biochemical Engineering/Biotechnology),Springer (2014). Compositions described herein may include a vector,such as a viral vector, e.g., a lentiviral vector, encoding arecombinant protein. In some embodiments, a vector, e.g., a viralvector, may comprise a nucleic acid encoding a recombinant protein.

Purification of protein therapeutics is described in Franks, ProteinBiotechnology: Isolation, Characterization, and Stabilization, HumanaPress (2013); and in Cutler, Protein Purification Protocols (Methods inMolecular Biology), Humana Press (2010).

Applications

By integrating coding genes into a RNA sequence template, the GeneWriter system can address therapeutic needs, for example, by providingexpression of a therapeutic transgene in individuals withloss-of-function mutations, by replacing gain-of-function mutations withnormal transgenes, by providing regulatory sequences to eliminategain-of-function mutation expression, and/or by controlling theexpression of operably linked genes, transgenes and systems thereof. Incertain embodiments, the RNA sequence template encodes a promotor regionspecific to the therapeutic needs of the host cell, for example a tissuespecific promotor or enhancer. In still other embodiments, a promotorcan be operably linked to a coding sequence.

In embodiments, the Gene Writer™ gene editor system can providetherapeutic transgenes expressing, e.g., replacement blood factors orreplacement enzymes, e.g., lysosomal enzymes. For example, thecompositions, systems and methods described herein are useful toexpress, in a target human genome, agalsidase alpha or beta fortreatment of Fabry Disease; imiglucerase, taliglucerase alfa,velaglucerase alfa, or alglucerase for Gaucher Disease; sebelipase alphafor lysosomal acid lipase deficiency (Wolman disease/CESD); laronidase,idursulfase, elosulfase alpha, or galsulfase for mucopolysaccharidoses;alglucosidase alpha for Pompe disease. For example, the compositions,systems and methods described herein are useful to express, in a targethuman genome factor I, II, V, VII, X, XI, XII or XIII for blood factordeficiencies.

In some embodiments, the heterologous object sequence encodes anintracellular protein (e.g., a cytoplasmic protein, a nuclear protein,an organellar protein such as a mitochondrial protein or lysosomalprotein, or a membrane protein). In some embodiments, the heterologousobject sequence encodes a membrane protein, e.g., a membrane proteinother than a CAR, and/or an endogenous human membrane protein. In someembodiments, the heterologous object sequence encodes an extracellularprotein. In some embodiments, the heterologous object sequence encodesan enzyme, a structural protein, a signaling protein, a regulatoryprotein, a transport protein, a sensory protein, a motor protein, adefense protein, or a storage protein.

Administration

The composition and systems described herein may be used in vitro or invivo. In some embodiments the system or components of the system aredelivered to cells (e.g., mammalian cells, e.g., human cells), e.g., invitro or in vivo. In some embodiments, the cells are eukaryotic cells,e.g., cells of a multicellular organism, e.g., an animal, e.g., a mammal(e.g., human, swine, bovine) a bird (e.g., poultry, such as chicken,turkey, or duck), or a fish. In some embodiments, the cells arenon-human animal cells (e.g., a laboratory animal, a livestock animal,or a companion animal). In some embodiments, the cell is a stem cell(e.g., a hematopoietic stem cell), a fibroblast, or a T cell. In someembodiments, the cell is a non-dividing cell, e.g., a non-dividingfibroblast or non-dividing T cell. In some embodiments, the cell is anHSC and p53 is not upregulated or is upregulated by less than 10%, 5%,2%, or 1%, e.g., as determined according to the method described inExample 30. The skilled artisan will understand that the components ofthe Gene Writer system may be delivered in the form of polypeptide,nucleic acid (e.g., DNA, RNA), and combinations thereof.

For instance, delivery can use any of the following combinations fordelivering the retrotransposase (e.g., as DNA encoding theretrotransposase protein, as RNA encoding the retrotransposase protein,or as the protein itself) and the template RNA (e.g., as DNA encodingthe RNA, or as RNA):

1. Retrotransposase DNA+template DNA

2. Retrotransposase RNA+template DNA

3. Retrotransposase DNA+template RNA

4. Retrotransposase RNA+template RNA

5. Retrotransposase protein+template DNA

6. Retrotransposase protein+template RNA

7. Retrotransposase virus+template virus

8. Retrotransposase virus+template DNA

9. Retrotransposase virus+template RNA

10. Retrotransposase DNA+template virus

11. Retrotransposase RNA+template virus

12. Retrotransposase protein+template virus

As indicated above, in some embodiments, the DNA or RNA that encodes theretrotransposase protein is delivered using a virus, and in someembodiments, the template RNA (or the DNA encoding the template RNA) isdelivered using a virus.

In one embodiments the system and/or components of the system aredelivered as nucleic acid. For example, the Gene Writer polypeptide maybe delivered in the form of a DNA or RNA encoding the polypeptide, andthe template RNA may be delivered in the form of RNA or itscomplementary DNA to be transcribed into RNA. In some embodiments thesystem or components of the system are delivered on 1, 2, 3, 4, or moredistinct nucleic acid molecules. In some embodiments the system orcomponents of the system are delivered as a combination of DNA and RNA.In some embodiments the system or components of the system are deliveredas a combination of DNA and protein. In some embodiments the system orcomponents of the system are delivered as a combination of RNA andprotein. In some embodiments the Gene Writer genome editor polypeptideis delivered as a protein.

In some embodiments the system or components of the system are deliveredto cells, e.g. mammalian cells or human cells, using a vector. Thevector may be, e.g., a plasmid or a virus. In some embodiments deliveryis in vivo, in vitro, ex vivo, or in situ. In some embodiments the virusis an adeno associated virus (AAV), a lentivirus, an adenovirus. In someembodiments the system or components of the system are delivered tocells with a viral-like particle or a virosome. In some embodiments thedelivery uses more than one virus, viral-like particle or virosome.

In one embodiment, the compositions and systems described herein can beformulated in liposomes or other similar vesicles. Liposomes arespherical vesicle structures composed of a uni- or multilamellar lipidbilayer surrounding internal aqueous compartments and a relativelyimpermeable outer lipophilic phospholipid bilayer. Liposomes may beanionic, neutral or cationic. Liposomes are biocompatible, nontoxic, candeliver both hydrophilic and lipophilic drug molecules, protect theircargo from degradation by plasma enzymes, and transport their loadacross biological membranes and the blood brain barrier (BBB) (see,e.g., Spuch and Navarro, Journal of Drug Delivery, vol. 2011, Article ID469679, 12 pages, 2011. doi:10.1155/2011/469679 for review).

Vesicles can be made from several different types of lipids; however,phospholipids are most commonly used to generate liposomes as drugcarriers. Methods for preparation of multilamellar vesicle lipids areknown in the art (see for example U.S. Pat. No. 6,693,086, the teachingsof which relating to multilamellar vesicle lipid preparation areincorporated herein by reference). Although vesicle formation can bespontaneous when a lipid film is mixed with an aqueous solution, it canalso be expedited by applying force in the form of shaking by using ahomogenizer, sonicator, or an extrusion apparatus (see, e.g., Spuch andNavarro, Journal of Drug Delivery, vol. 2011, Article ID 469679, 12pages, 2011. doi:10.1155/2011/469679 for review). Extruded lipids can beprepared by extruding through filters of decreasing size, as describedin Templeton et al., Nature Biotech, 15:647-652, 1997, the teachings ofwhich relating to extruded lipid preparation are incorporated herein byreference.

Lipid nanoparticles are another example of a carrier that provides abiocompatible and biodegradable delivery system for the pharmaceuticalcompositions described herein. Nanostructured lipid carriers (NLCs) aremodified solid lipid nanoparticles (SLNs) that retain thecharacteristics of the SLN, improve drug stability and loading capacity,and prevent drug leakage. Polymer nanoparticles (PNPs) are an importantcomponent of drug delivery. These nanoparticles can effectively directdrug delivery to specific targets and improve drug stability andcontrolled drug release. Lipid-polymer nanoparticles (PLNs), a new typeof carrier that combines liposomes and polymers, may also be employed.These nanoparticles possess the complementary advantages of PNPs andliposomes. A PLN is composed of a core-shell structure; the polymer coreprovides a stable structure, and the phospholipid shell offers goodbiocompatibility. As such, the two components increase the drugencapsulation efficiency rate, facilitate surface modification, andprevent leakage of water-soluble drugs. For a review, see, e.g., Li etal. 2017, Nanomaterials 7, 122; doi:10.3390/nano7060122.

Exosomes can also be used as drug delivery vehicles for the compositionsand systems described herein. For a review, see Ha et al. July 2016.Acta Pharmaceutica Sinica B. Volume 6, Issue 4, Pages 287-296;https://doi.org/10.1016/j.apsb.2016.02.001.

A Gene Writer system can be introduced into cells, tissues andmulticellular organisms. In some embodiments the system or components ofthe system are delivered to the cells via mechanical means or physicalmeans.

Formulation of protein therapeutics is described in Meyer (Ed.),Therapeutic Protein Drug Products: Practical Approaches to formulationin the Laboratory, Manufacturing, and the Clinic, Woodhead PublishingSeries (2012).

All publications, patent applications, patents, and other publicationsand references (e.g., sequence database reference numbers) cited hereinare incorporated by reference in their entirety. For example, allGenBank, Unigene, and Entrez sequences referred to herein, e.g., in anyTable herein, are incorporated by reference. Unless otherwise specified,the sequence accession numbers specified herein, including in any Tableherein, refer to the database entries current as of Aug. 27, 2018. Whenone gene or protein references a plurality of sequence accessionnumbers, all of the sequence variants are encompassed.

EXAMPLES

The invention is further illustrated by the following examples. Theexamples are provided for illustrative purposes only and are not to beconstrued as limiting the scope or content of the invention in any way.

Example 1: Delivery of a Gene Writer™ System to Mammalian Cells

This example describes a Gene Writer™ genome editing system delivered toa mammalian cell for site-specific insertion of exogenous DNA into amammalian cell genome.

In this example, the polypeptide component of the Gene Writer™ system isthe R2Bm protein from Bombyx mori and the template RNA component is RNAfor the R2Bm retrotransposase from Bombyx mori containing a mutation inthe reverse transcriptase domain that renders the retrotransposaseinactive.

HEK293T cells are transfected with the following test agents:

1. Scrambled RNA control2. RNA coding for the polypeptide described above3. Template RNA described above

4. Combination of 2 and 3

After transfection, HEK293T cells are cultured for at least 4 days andthen assayed for site-specific genome editing. Genomic DNA is isolatedfrom each group of HEK293 cells. PCR is conducted with primers thatflank the R2Bm integration site in 28s rRNA genes. The PCR product isrun on an agarose gel to measure the length of the amplified DNA.

A PCR product of the expected length, indicative of a successful GeneWriting™ genome editing event that inserts the sequence for the mutatedR2Bm retrotransposase into the target genome, is observed only in cellsthat were transfected with the complete Gene Writer™ system of group 4above.

Example 2: Site-Specific Targeted Delivery of a Gene Writer™ System intoInsect Cells

This example describes a Gene Writer™ genome editing system delivered toan insect cell at a specific target site of the genome.

In this example, the polypeptide component of the Gene Writer™ system isderived from R2Bm of Bombyx mori, which is modified by replacing its DNAbinding domain in the amino terminus of the polypeptide with aheterologous zinc-finger DNA binding domain. The zinc finger DNA bindingdomain is known to bind to DNA in the BmBLOS2 loci of B. mori cells(Takasu et al., insect Biochemistry and Molecular Biology 40(10):759-765, 2010). The template RNA is RNA for the R2Bm retrotransposasefrom Bombyx mori containing a mutation in the reverse transcriptasedomain that renders the retrotransposase inactive. Furthermore, thetemplate RNA is modified at the 5′ end to have 180 bases of homology tothe target DNA site.

B. mori insect cell lines are transfected with the following testagents:

1. Scrambled RNA control2. RNA coding for polypeptide component described above3. Template RNA described above

4. Combination of 2 and 3

After transfection, the cells are cultured for at least 4 days andassayed for site-specific Gene Writing™ genome editing. Genomic DNA isisolated from the cells and PCR is conducted with primers that flank thetarget integration site in the genome. The PCR product is run on anagarose gel to measure the length of DNA. A PCR product of the expectedlength, indicative of a successful Gene Writing™ genome editing eventthat inserts the sequence for the mutated R2Bm retrotransposase into thetarget insect cell genome, is observed only in cells that weretransfected with the complete Gene Writer™ system of group 4 above.

Example 3: Site-Specific Targeted Delivery of a Gene Writer™ System intoMammalian Cells

This example describes a Gene Writer™ genome editor system used toinsert a heterologous sequence into a specific site of the mammaliangenome.

In this example, the polypeptide of the system is the R2Bm protein fromBombyx mori and the template RNA component is RNA coding for the GFPprotein and flanked at the 5′ end by the 5′ UTR and at the 3′ end by the3′ UTR of the R2Bm retrotransposase from Bombyx mori. The GFP gene hasan internal ribosomal entry site upstream of its start codon and a polyAtail downstream of its stop codon.

HEK293 cells are transfected with the following test agents:

1. Scrambled RNA control2. RNA coding for the polypeptide described above3. Template RNA coding for GFP described above

4. Combination of 2 and 3

After transfection, HEK293 cells are cultured for at least 4 days andthen assayed for a site-specific Gene Writing™ genome editing event.Genomic DNA is isolated from the HEK293 cells and PCR is conducted withprimers that flank the R2Bm integration site in 28s rRNA genes. The PCRproduct is run on an agarose gel to measure the length of DNA. A PCRproduct of the expected length, indicative of a successful Gene Writing™genome editing event, is detected in cells transfected with the testagent of group 4 (complete Gene Writer™ system). This resultdemonstrates that a Gene Writing genome editing system can insert anovel transgene into the mammalian cell genome.

The transfected cells are cultured for a further 10 days, and aftermultiple cell culture passages are assayed for GFP expression via flowcytometry. The percent of cells that are GFP positive from each cellpopulation are calculated. GFP positive cells are detected in thepopulation of HEK293 cells that were transfected with the test agent ofgroup 4 (complete Gene Writer™ system). This result demonstrates thatthe novel transgene written into the mammalian cell genome is expressed.

Example 4: Targeted Delivery of a Gene Expression Unit into MammalianCells Using a Gene Writer™ System

This example describes the making and using of a Gene Writer genomeeditor to insert a heterologous gene expression unit into the mammaliangenome.

In this example, the polypeptide of the Gene Writer system is derivedfrom the R2Bm polypeptide of Bombyx mori as modified by replacing itsDNA binding domain in the amino terminus of the polypeptide with aheterologous zinc-finger DNA binding domain. The zinc finger DNA bindingdomain is known to bind to DNA in the AAVS1 locus of human cells(Hockemeyer et al., Nature Biotechnology 27(9): 851-857, 2009). Thetemplate RNA comprises a gene expression unit. A gene expression unitcomprises at least one regulatory sequence operably linked to at leastone coding sequence. In this example, the regulatory sequences includethe CMV promoter and enhancer, an enhanced translation element, and aWPRE. The coding sequence is the GFP open reading frame. The geneexpression unit is flanked at the 5′ end by 180 bases of homology to thetarget DNA site and at the 3′ end by the 3′ UTR of the R2Bmretrotransposase from Bombyx mori.

HEK293 cells are transfected with the following test agents:

1. Scrambled control RNA2. RNA coding for the polypeptide component described above3. Template RNA comprising the gene expression unit (as described above)4. The complete Gene Writer system comprising both (2) and (3)

After transfection, HEK293 cells are cultured for at least 4 days andassayed for site-specific Gene Writing genome editing. Genomic DNA isisolated from the HEK293 cells and PCR is conducted with primers thatflank the target integration site in the genome. The PCR product is runon an agarose gel to measure the length of DNA. A PCR product of theexpected length, indicative of a successful Gene Writing™ genome editingevent, is detected in cells transfected with the test agent of group 4(complete Gene Writer™ system).

The transfected cells are cultured for a further 10 days, and aftermultiple cell culture passages are assayed for GFP expression via flowcytometry. The percent of cells that are GFP positive from each cellpopulation are calculated. GFP positive cells are detected in thepopulation of HEK293 cells that were transfected with group 4 testagent, demonstrating that a gene expression unit added into themammalian cell genome via Gene Writing genome editing is expressed.

Example 5: Targeted Delivery of a Gene Expression Unit into an IntronicRegion of Mammalian Cells Using a Gene Writer™ System

This example describes the making and use of a Gene Writing genomeediting system to add a heterologous sequence into an intronic region toact as a splice acceptor for an upstream exon.

The target integration site is the first intron of the albumin locus.Splicing into the first intron a new exon containing a splice acceptorsite at the 5′ end and a polyA tail at the 3′ end will result in amature mRNA containing the first natural exon of the albumin locusspliced to the new exon. Because the first exon of albumin is removedupon protein processing, the cell expressing the newly formed gene unitwill secrete a mature protein comprising only the new exon.

In this example, the Gene Writer genome editor polypeptide is derivedfrom the R2Bm Gene Writer genome editor of Bombyx mori as modified byreplacing the DNA binding domain in the amino terminus of thepolypeptide with a heterologous zinc-finger DNA binding domain. The zincfinger DNA binding domain is known to bind tightly to the albumin locusin the first intron as described in Sarma et al., Blood 126, 15:1777-1784, 2015. The template RNA is RNA coding for EPO with a spliceacceptor site immediately 5′ to the first amino acid of mature EPO (thestart codon and signal peptide is removed) and a 3′ polyA taildownstream of the stop codon. The EPO RNA is further flanked at the 5′end by 180 bases of homology to target DNA site and at the 3′ end by the3′ UTR of the R2Bm retrotransposase from Bombyx mori.

HEK293 cells are transfected with the following test agents:

1. Scrambled control RNA2. RNA coding for the polypeptide described above3. Template RNA comprising the EPO splice acceptor described above4. The complete Gene Writer system comprising both (2) and (3)

After transfection, HEK293 cells are cultured for at least 4 days andassayed for site-specific Gene Writing genome editing and appropriatemRNA processing. Genomic DNA is isolated from the HEK293 cells. Reversetranscription-PCR is conducted to measure the mature mRNA containing thefirst natural exon of the albumin locus and the new exon. The RT-PCRreaction is conducted with forward primers that bind to the firstnatural exon of the albumin locus and with reverse primers that bind toEPO. The RT-PCR product is run on an agarose gel to measure the lengthof DNA. A PCR product of the expected length is detected in cellstransfected with the test agent of group 4, indicative of a successfulGene Writing genome editing event and a successful splice event. Thisresult demonstrates that a Gene Writing genome editing system can add aheterologous sequence encoding a gene into an intronic region to act asa splice acceptor for the upstream exon.

The transfected cells are cultured for a further 10 days, and aftermultiple cell culture passages are assayed for EPO secretion in the cellsupernatant. The amount of EPO in the supernatant is measured via an EPOELISA kit. EPO is detected in HEK293 cells that were transfected withthe test agent of group 4, demonstrating that a heterologous sequencecan be added into an intronic region via Gene Writing genome editing, toact as a splice acceptor for the upstream exon and is activelyexpressed.

Example 6: Targeted Delivery of R2Tg Retrotransposon to Mammalian Cells

This example describes targeted integration of the R2Tg retrotransposonelement (see first row of Table 3 herein) to mammalian cells via DNA orRNA delivery.

R2Tg is an endogenous retrotransposon from the zebra finch (Taenopygiaguttata). Because non-LTR R2 elements are not present in the humangenome and are thought to be highly site-specific, the ability of R2Tgto accurately and efficiently integrate itself into the human genomewould demonstrate the capability to perform genomic targeted integrationand possibly enable human gene therapy.

In the DNA delivery method, plasmid harboring R2Tg (PLV014) was designedand synthesized such that the R2Tg element was codon optimized andflanked by its native un-translated regions (UTRs), with or withoutfurther flanking by 100 bp homology to the rDNA target locus. The R2Tgelement expression was driven by the mammalian CMV promoter. Further, a1 bp deletion mutant (678*) having a frameshift in the coding sequenceof the retrotransposase was constructed as an inactivated control(“frameshift mutant”). Each plasmid was introduced into HEK393T cellsvia FuGENE® HD transfection reagent. HEK293T cells were seeded in96-well plate, 10,000 cells/well 24 hr before transfection. On thetransfection day, 0.5 μl transfection reagent and 80 ng DNA was mixed in100 Opti-MEM and incubated for 15 min at room temperature. Then thetransfection mixture was added to the medium of the seeded cells. 3 daysafter transfection, genomic DNA was extracted for retrotranspositionassays.

Next, integration of the R2Tg transposase into the human genome wasassessed. Based on homology to the finch genome, a putative integrationsite in human rDNA was tested. Advanced Miseq and ddPCR assays were usedto assess integration.

Bias in Miseq library construction was eliminated by introducing randomunique molecular indices (UMIs) into initial PCRs (FIG. 7). Nested PCRwas performed by first amplifying the expected 3′ junction of R2Tg andthe rDNA locus for 30 cycles. One Miseq adapter, a multiplexing barcode,and an 8 bp UMI were introduced at this step. A second PCR was used tofurther enrich for expected products and add the second Miseq adapter.Samples were sequenced on the Miseq for 300 cycles. Afterdemultiplexing, the samples were analyzed via Matlab. First, the UMI oneach sequence was located by searching for neighboring sequence. Adatabase of UMIs was created and next collapsed by uniqueness. For eachunique read, for a search was performed for the sequence of the expectedrDNA integration site and isolated sequences of aligned human genomicDNA and exogenous DNA. Exogenous DNA was then aligned to the expectedintegration sequence. Results of the Miseq analysis pipeline are shownin FIGS. 8A-8B. Extensive unique integrations into the predictedintegration site were found in cells treated with the wildtype R2Tgconstruct flanked by 100 bp homology to the target rDNA locus, but notwith the frameshift mutant controls. Most integration events have acomplete template RNA sequence integrated in the 565 bp most proximal tothe integration site as demonstrated by sequencing reads that alignperfectly to the expected sequence. A subset of integration events withthe experimental R2Tg have either a ˜300 bp or ˜450 truncation as basedupon sequencing reads that align to the expected sequence after a gapdirectly adjacent to the target site (FIG. 8A). More specifically,86.17% of integrants observed were non-truncated in the 565 bp mostproximal to the integration site. In contrast, FIG. 8B shows nointegration events detected. Constructs without flanking rDNA homologyshowed insignificant integration signals near noise.

ddPCR was next performed to confirm integration and assess integrationefficiency. A Taqman probe was designed to the 3′UTR portion of the R2Tgelement. A forward primer was synthesized to bind directly upstream ofthe probe, and a reverse primer was synthesized to bind the rDNA.Therefore, amplification of the expected product across the integrationjunction degrades the probe and creates a fluorescent signal. ddPCR wasperformed on several replicate experiments of the above plasmids todetermine the average copy number of the R2Tg integration event. Theresults of ddPCR copy number analysis (in comparison to reference geneRPP30) are shown in FIG. 9. Across several plasmid transfectionconditions, average integration of 5 or more copies of R2Tg per genomeat the target site when delivered with homology was noted withsignificant increase above control constructs. In contrast, the averagecopy number per genome in the frameshift mutant negative control wastypically lower than 1. Insignificant signal was seen when constructswithout homology were delivered to cells. The experiments collectivelysuggest efficient integration of the R2Tg retrotransposon into humancells at the target site.

In the RNA delivery method, R2Tg RNA (RNAV019) was designed such thatthe R2Tg element was codon optimized and flanked by its nativeuntranslated regions (UTRs). More specifically, the construct includes,in order: a T7 promoter, a 5′ 28S target homology region 100 nucleotidesin length, a R2Tg wild-type 5′ UTR, the R2Tg codon-optimized codingsequence, a R2Tg wild type 3′ UTR, and a 3′ 28S target homology region100 nucleotides in length. The 100 bp 28S homology sequences were addedoutside of the UTRs to enhance the integration. R2Tg RNA wassynthesized, and cap and polyA tail were added. The R2Tg elementtranscription was driven by the T7 promoter. The RNA was introduced intoHEK393T cells via Lipofectamine™ RNAiMAX or TransIT®-mRNA transfectionreagent with series of RNA dosages. HEK293T cells were seeded in 96-wellplate 24 hr before transfection. On the transfection day, transfectionreagent and RNA were mixed in 10 μl Opti-MEM, and the transfectionmixture was added to the medium of the seeded cells. 3 days aftertransfection, genomic DNA was extracted to measure retrotranspositionefficiency using ddPCR with the same design as the DNA delivery.

The results of ddPCR copy number analysis (normalized to reference geneRPP30) are shown in FIG. 12. Across several transfection conditions, theaverage integration was measured to be 0.01 of R2Tg copies per genome,significantly above the limit of detection. The results indicatesuccessful integration of the R2Tg retrotransposon into human cellsusing an RNA delivery method.

Example 7: Targeted Delivery of a Heterologous Object Sequence UsingR2Tg Retrotransposon to Mammalian Cells

This example describes the delivery of a transgene to human cells byutilizing the R2Tg retrotransposon system with multiple deliverymachineries, including RNA-mediated delivery of a heterologous objectsequence to human cells by utilizing the R2Tg retrotransposon system.

R2 proteins recognize their template RNA structure in untranslatedregions (UTRs) of each element to form ribonucleoprotein particles,which serve as the intermediates of downstream integration into a hostgenome. Therefore, the decoupling of UTRs from their native context andthe introduction of UTRs into alternate exogenous sequence wasengineered to deliver into the genome a desired nucleic acid using R2Tgmachinery.

Trans-transgene integration was tested by constructing 1) R2Tg codingsequence and 2) transgene cassette flanked by R2Tg UTR sequences and 100bp homology to 28S rDNA into separate driver and transgene plasmids,respectively. FIG. 13 illustrates the dual plasmid system. The dualplasmids were introduced into HEK293T cells via FuGENE® HD transfectionreagent at multiple driver to transgene molar ratios. In addition to theWT R2Tg driver, backbone plasmid was used as a control. HEK293T cellswere seeded in 96-well plates at 10,000 cells/well 24 hr beforetransfection. On the transfection day, transfection reagent and plasmidswere mixed in 10 μl Opti-MEM and incubated for 15 minutes at roomtemperature, then added to the medium of the seeded cells. 3 days aftertransfection, genomic DNA was extracted for ddPCR assays to investigatethe trans-retrotransposition efficiency. FIG. 14 demonstrates the ddPCRresults for conditions with excess of transgene relative to driver.

Similar to the trans-transgene delivery with plasmids, RNA delivery wasperformed by constructing an amplicon of the coding sequence of R2Tgpreceded by the T7 promoter sequence. The constructed ampliconscontained the experimental R2Tg element as well as the 1 bp deletionframeshift mutant control. Separately, an amplicon was constructed thatcontained exogenous sequence coding for GFP and an EGF1-alpha reporterthat was flanked regions sufficient to drive integration into the genomeby R2Tg. More specifically, the construct included: a T7 promoterdriving transcription of the RNA, wherein the RNA comprises, from 5′ to3′, (a) a 5′ 28S homology region of 10 nt in length, (b) a 5′untranslated region, (c) an anti-sense TKpA polyA sequence, (d) ananti-sense heterologous object sequence that encodes GFP, (e) ananti-sense Kozak sequence, (f) an anti-sense EF1 alpha promoter, (g) a3′ untranslated region that binds the GeneWriter protein, and (h) a 3′28S homology region of 10 nt in length. Each RNA was transcribed via theNew England Biolabs HiScribe T7 ARCA kit and purified via Zymo RNA cleanand concentrator.

The resulting heterologous object RNA and R2Tg RNA (either theexperiment R2Tg element or frameshift mutant) were introduced into humanHEK293T cells via TransIT®-mRNA Transfection Kit at 1:1 molar ratio.HEK293T cells were seeded in 96-well plate, 40,000 cells/well 24 hrbefore transfection. On the transfection day, 10 transfection reagentand 500 ng total RNA was mixed in 10 μl Opti-MEM and incubated for 5 minat room temperature. Then the transfection mixture was added to themedium of the seeded cells. 3 days after transfection, genomic DNA wasextracted for PCR assays.

Nested PCR was performed by with a first 30 rounds of PCR across the 3′end of the expected transgene-rDNA junction, followed by 20 additionalrounds of PCR amplification using an inner primer set. One of threereplicates of nested PCR performed on genomic DNA extracted from cellstreated with the wild-type transposase reaction produced a PCR productof the expected size (approximately 596 bp). In contrast, no PCR productwas observed in genomic DNA extracted from cells treated with theframeshift-inactivated R2Tg mutant control, or no-transfection control.The PCR product was gel-purified via Zero Blunt® TOPO® PCR Cloning Kit,and the resulting colonies were Sanger sequenced. Each individual PCRproduct sequence was then aligned to the expected integration sequence.The fraction of PCR product sequences that align to the expectedintegrated heterologous object sequence is shown in FIG. 10. Themajority of PCR products had the expected integrant as demonstrated bythe sequencing alignment directly adjacent to the expected integrationsite at the right side of the alignment figure. This demonstratesRNA-mediated integration of the exogenous sequence via R2Tg machineryinto human cells.

Example 8: Targeted Delivery of R2Tg Retrotransposon to Mammalian Cells

This example describes targeted integration of the R2Tg retrotransposonelement to mammalian cells via DNA delivery.

Plasmid harboring R2Tg (PLV014) and control plasmid were designed andsynthesized as described above in Example 6. Each plasmid was introducedinto HEK393T cells via FuGENE® HD transfection reagent. HEK293T cellswere seeded in 96-well plate, 10,000 cells/well 24 hr beforetransfection. On the transfection day, 0.5 μl transfection reagent and80 ng DNA was mixed in 10 μl Opti-MEM and incubated for 15 min at roomtemperature. Then the transfection mixture was added to the medium ofthe seeded cells. 3 days after transfection, genomic DNA was extractedfor retrotransposition assays or cells were frozen and underwenttargeted locus amplification.

Target locus amplification was performed against hg38 reference humangenome and the rDNA locus sequence hsu13369 (GenBank: U13369.1). Twoindependent primer sets were used to perform targeted locusamplification. Analysis with both primer sets showed that the 28S rDNAlocus sequence is the only integration site detected above a 1%threshold. Thus, integration of the R2Tg transposon in mammalian cellsis specific to this target site.

Example 9: Improved Trans RNA-Templated Integration into Mammalian Cellsby RNA Refolding or Ratio of Driver to Template RNA

RNA templates are designed as in previous examples. Two RNAs consistingof a driver and a transgene payload are delivered to mammalian cells. Tobetter promote folding, denaturing the payload RNA by heating to 95 Cand cooling to room temperature are performed to encourage propersecondary structure formation. In some embodiments, cooling the RNA toroom temperate will increase integration efficiency.

The molar ratio of transgene to driver is also varied to evaluatesuitable stoichiometry of components. Integration is analyzed via ddPCRand sequencing. In some embodiments, a higher ratio of driver totransgene is used. In some embodiments, a higher ratio of transgene todriver is used.

Previous examples with cis transgene integration are similarly assayedfor stoichiometry of driver to payload. Integration is analyzed viaddPCR and sequencing. In some embodiments, a higher ratio of drivertranscription or translation to transgene transcription will result inhigher integration efficiency. In some embodiments, a higher ratio oftransgene transcription to driver transcription and translation willresult in higher integration efficiency.

Example 10: Hybrid Capture Assay

A hybrid capture experiment was performed to obtain an unbiased view ofthe specificity of retrotransposon integration into a target site.Retrotransposon experiments were performed as in previous examples byintegrating R2Tg flanked by its native UTRs and 100 bp of homology toeither side of the expected R2 rDNA target. The rDNA target site had twoflanking sets of 100 nucleotides identity to the corresponding nativetarget site. The retrotransposon was delivered to human 293T cells viaplasmid or mRNA. Genomic DNA was extracted after 72 hours. Afterextraction, each genomic DNA sample was subjected to hybrid captureaccording to protocol with a custom probe set (Twist). Biotinylatedprobes were designed such that ˜120 bp probes spanned both strands ofthe R2Tg coding sequence and UTRs. First, a next-generation library wascreated by fragmentation of the genomic DNA and ligation of sequencingadapters according to a protocol from Twist (available on the world wideweb at: twistbioscience.com/ngs_protocol_custompanel_hybridcap). Next,probes were hybridized to genomic DNA libraries and the enriched sampleswere amplified. Final libraries were sequenced on the Miseq using 300 bppaired-end reads. Custom Matlab scripts were used to analyze reads. Theresulting analysis is shown in FIGS. 15A and 15B for RNA delivery.Hybrid capture indicated on-target integration of R2Tg to the expectedlocus. With RNA delivery, 1 possible off-target with a single read wasidentified at an unexpected 3′ junction in the data, compared to morethan 100 reads at the expected locus, indicating a specificity ofgreater than 100:1. At the 5′ junction, all 50 reads were at theexpected locus, indicating a specificity of greater than 50:1. Thisexperiment indicates a high specificity of integration.

Example 11: Long-Read PacBio Analysis

Long-range PCR amplification can be performed to measure integration ofthe desired full-length sequence into the target site in the humangenome and to measure whether mutations are introduced during insertion.Retrotransposon integration experiments are performed as described inprevious examples. In one example, PCR amplification is used to generateamplicons by designing one primer targeting the genomic integration siteand one primer targeting the integrant sequence. In this example, theseprimers are designed to maximize the length of the amplified genomiclocus fused with the integrant sequence. By pooling amplicons spanningboth ends of the integrant and performing long-read next-generationsequencing, the fidelity of each integration is be evaluated.

In another example, hybrid capture is performed as described in aprevious example but with a larger target library length during initiallibrary generation. The resulting library is then subjected to long-readnext-generation sequencing.

In some embodiments, long-read next generation sequencing will show thatthere are less than 10%, 5%, 2%, 1%, 0.5%, 0.2%, or 0.1% SNPs in theintegrated DNA across samples. In some embodiments, long-read nextgeneration sequencing will show that less than 10%, 5%, 2%, or 1% ofintegrated DNA has a SNP. In some embodiments, long-read next generationsequencing will show that less than 10%, 5%, 2%, or 1% of integrated DNAhas an internal deletion. In some embodiments, long-read next generationsequencing will show that less than 10%, 5%, 2%, 1%, 0.5%, 0.2%, or 0.1%of total integrated DNA across the population is deleted. In someembodiments, long-read next generation sequencing will show that lessthan 10%, 5%, 2%, or 1% of integrated DNA is truncated.

Example 12: Experiment with Different Homology Lengths and PointMutations in Homology

In this example, experiments are designed to characterize suitablelengths and starting positions of homology to the target site forefficient retrotransposon integration. Also, the homology is used tosupport the mechanism of integration being reverse transcription-driven.

A series of SNPs were introduced within the 100 bp downstream homologyof R2Tg plasmids by modifying plasmid PLV014. The design of the SNPs islisted in FIG. 16. After the transfection, nested PCR was applied torecover the 3′ integration junction site, producing a PCR product withan expected amplicon size of about 738 bp, and the PCR product wasSanger sequenced to check whether any SNPs were incorporated. In thisexperiment, a lack of SNP genetic markers being incorporated into thejunction sequences indicates that the integration was driven by reversetranscription. The SNP design and the sequencing result are illustratedin FIG. 16. No SNP introduction was observed for the 18 genetic markersdesigned, consistent with the integration of R2Tg being directed byreverse transcription.

This example also describes the evaluation different homology regions tothe target site to identify shorter regions that promote efficientintegration into the genome. This example describes two approaches.First, different windows of 100 bp of homology to the target site aretested, starting from bp 1-100 3′ of the target site, then testing 2-1013′ of the target site, 3-103 3′ of the target site, and so on, throughbp 30-131 3′ of the target site. Second, shorter lengths of homology tothe target site sufficient for DNA integration are tested, starting withbp 0-100 3′ of the target site, then testing 0-95 3′ of the target site,0-90 3′ of the target site, etc. through bp 0-10 3′ of the target site.After the transfection of each plasmid into 293T cells, ddPCR is used tomeasure the retrotransposition efficiency.

In this example, different UTR regions with different lengths areevaluated to identify shorter sequences for efficient integration intothe genome. The 3′UTR is tested by dividing this 325 bp sequence into 3regions, 1-100 bp, 101-200 bp, and 201-325 bp. Constructs of R2Tgcontaining each truncated 3′UTR are generated to test the integrationefficiency respectively.

Example 13: Assess Whether p53 or Other Repair Pathways are Upregulated

This example describes an evaluation of the effect of exogenous R2Tgretrotranspositon on gene expression, especially tumor suppressor andDNA repair genes. An R2Tg expressing plasmid is delivered to multiplecancer cell lines, including 293T, MCF-7, and T47D. After confirmationof integration in each cell line, RNA-seq is conducted to assess theeffect on gene expression profile. Gene set enrichment analysis is thenapplied to evaluate whether any DNA repair pathways are upregulatedafter retrotransposition. MCF-7 and T47D are breast cancer cell lineswith wild type and mutant p53 respectively, which are be used toevaluate the relationship between p53 and retrotranspositonspecifically. In some embodiments, p53 is not upregulated when aretrotransposon Gene Writer integrates into the genome. In someembodiments, no DNA repair genes are upregulated when a retrotransposonGene Writer integrates into the genome. In some embodiments, no tumorsuppressor genes are upregulated when a retrotransposon Gene Writerintegrates into the genome.

Example 14: Retrotransposition in Presence of DNA Repair Inhibitors

In this example, experiments will test the effect of different DNArepair pathways on R2Tg retrotransposition via the application of DNArepair pathway inhibitors or DNA repair pathway deficient cell lines.When applying DNA repair pathway inhibitors, PrestoBlue cell viabilityassay is performed first to determine the toxicity of the inhibitors andwhether any normalization should be applied for following assays. SCR7is an inhibitor for NHEJ, which is applied at a series of dilutionsduring R2Tg delivery. PARP protein is a nuclear enzyme that binds ashomodimers to both single- and double-strand breaks. Thus, itsinhibitors are be used in the test of relevant DNA repair pathways,including homologous recombination repair pathway and base excisionrepair pathway. The experiment procedure is the same with that of SCR7.Cell lines with deficient core proteins of nucleotide excision repair(NER) pathway are used to test the effect of NER on R2Tgretrotransposition. After the delivery of R2Tg element into the cell,ddPCR is be used to evaluate the retrotransposition in the context ofinhibition of DNA repair pathways. Sequencing analysis is also beperformed to evaluate whether certain DNA repair pathway plays a role inthe alteration of integration junction. In some embodiments, R2Tgintegration into the genome will not be decreased by the knockdown ofany DNA repair pathways, suggesting that R2Tg does not rely on the hostcell pathways for DNA integration.

Example 15: Retrotransposition in Fibroblasts and in T Cells

In this example, the previously performed R2Tg retrotranspositionanalysis of 293T cells is repeated in non-dividing cells, includingfibroblast and T cells. Compared to 293T cells, non-dividing cells aresometimes more difficult to transfect with lipid reagent. Thus,nucleofection is used for the delivery of R2Tg element. The subsequentretrotransposition assay for integrating efficiency and sequencinganalysis will be performed as described herein for 293T cells. In someembodiments, R2Tg integrates into the genome of fibroblasts and T cells.

Example 16: Single Cell ddPCR

In this example, a quantitative assay is used to determine the frequencyof targeted genome integration at single cell level, and thatinformation can be compared to the copy number of targeted genomeintegration per genome quantified from genomic DNA.

Approximately 5000 transfected cells will be collected and mixed withddPCR reaction mixture before distributing into about 20,000 droplets,with the aim of each droplet containing one cell or no cells. ddPCRassays including 5′UTR and 3′UTR assays will be performed as describedabove to determine the frequency of R2 or transgene integration atsingle cell level. A control experiment will be performed in parallelusing genomic DNA harvested from the same number of cells to determinethe targeted genome integration efficiency per genome. In someembodiments, the frequency of targeted genome integration at the singlecell level is calculated to be 1-80%, e.g., 25%, wherein the indicatedpercentage of cells have one or more copies of the transgene integratedinto the desired locus.

Example 17: Single Cell Analysis Via Colony Isolation

In this example, a quantitative assay is used to determine genomeintegration copy number in cell colonies derived from single cell.

Single cell colonies will be isolated by colony picking up or by limiteddilution and cultured in a 96 well format. When the cells reach >80%confluency, half of the cells will be frozen for backup and genomic DNAfrom the other half of the cells will be harvested for ddPCR. OptimizedddPCR assays including 5′UTR and 3′UTR assays will be performed asdescribed previously to determine the frequency of R2 or transgeneintegration. At least 96 colonies will be screened for each R2 elementwith appropriate controls. The total number of colonies to be screenedwill be determined by single cell ddPCR data if applicable or the firstset of single cell colony screen data. In some embodiments, thefrequency of targeted genome integration at the single cell level willbe calculated to be 1-80%, e.g., 25%, wherein the indicated percentageof cells have a single copy of the transgene integrated into the desiredlocus. The assay can also be used to determine the percentage ofcolonies that have more than one copy of the transgene integrated intothe desired locus.

Example 18: DNA Binding Affinity and/or Re-Targeting

The DNA targeting module of wild-type R2 is made of a cysteine-histidinezinc finger and c-Myb transcription factor binding motifs. ThisN-terminal module can be substituted with different DNA binding modulessuch as DNA binding protein(s) (e.g., transcription factors), zincfinger(s) (e.g., natural or designed motifs), and/or nucleic acidguided, catalytically inactive endonucleases (e.g., Cas9 bound with aguide RNA (e.g., sgRNA) to form a Cas9-RNP). This DNA binding module isswapped for the naturally occurring module and, in some cases placedwith a flexible linker attaching it to the RNA binding/RT module.Additionally, in some constructions, this new DNA binding module isplaced in tandem with the same and/or different DNA binding modules.Furthermore, some constructions may split the GeneWriter protein whereone protein molecule contains the RNA binding module and the otherprotein contains the RT and endonuclease modules. In some embodiments,swapping of the DNA module increases specificity and/or affinity to agenomic location and in some cases allows for the specific targeting ofnew genomic locations.

Example 19: Assays to Measure DNA Binding Affinity

DNA binding activity of GeneWriters described herein (and DNA bindingdomains for the same) can be tested, e.g., as described in this example.DNA binding modules are purified by recombinantly expressing them incells (e.g., E. coli) or they are expressed in a cell-free reactions oftranscription and translation (e.g., T7 RNA polymerase+wheat germextract). The purified DNA binding module(s) is tested for bindingaffinity by measuring the Kd in a binding assay (e.g., EMSA,Fluorescence anisotropy, dual-filter binding, FRET, SPR, orthermophoresis (temperature related intensity change). The protein (DNAbinding module) is labeled and/or the DNA molecule is labeled with amolecule that is compatible with the above binding assays (e.g., dye,radioisotope (for example, Protein: ³⁵S-methionine, maleimide dye, DNA:³²P end or internal label, DNA with a linked amine reacted withNHS-ester dye). The molecules are measured by changing theirconcentrations and fitting to a binding curve which calculates thebinding affinity. In some assays, the nucleic acid sequence specificityis tested by mutational analysis of the DNA sequence or mutation to theDNA binding module by amino acid changes or alterations toprotein-nucleic acid complex (e.g., Cas9-RNP DNA binding module). Insome embodiments, increasing the Kd of the DNA binding module willdecrease off-target insertions and, in some cases, will increase theactivity of on-target sites by increasing the dwell time of the R2-RNAcomplex at the specific genomic location.

Example 20: Assays to Determine Global Specificity De Novo

The DNA binding module is expressed in cells (e.g., animal cells, e.g.,human cells) as the DNA binding module alone, in the context of thefull-length retrotransposon R2, or a control without retrotransposase.The expression of the module or retrotransposon is delivered to cellsusing conventional methods of delivering DNA, RNA, or protein. Thecomplex is crosslinked (e.g., using chemical or UV light) or is notcrosslinked. The cells are lysed and treated with DNase I so that onlythe bound DNA is protected from degradation. DNA is extracted, NGSlibrary preparation of DNA fragments and de novo binding sites areidentified, analogous to ChIP-seq or DIG-seq. In some embodiments,potential off-target sites are identified that can be followed-up toremove false-positives. In other embodiments this assay confirms the invitro assay on the specificity of the DNA binding module to bind at itsintended site and not at others.

An orthogonal assay to identify DNA binding sites in high-throughputuses the method described by Boyle et al, PNAS 2017 where the DNAbinding domain is tested in a cell-free setting to determine specificityalong with systematic analysis of sequence mutants related to the newDNA binding module.

Example 21: Modularity of RNA Molecule

The RNA molecule binds to the R2 protein via interactions found in thereverse transcriptase module, designated as a sub-module “RNA binding”.The protein recognizes specific structures in the 5′ and/or 3′ UTRs tointeract with the RNA. In some embodiments, swapping of the UTR modulesincreases protein interactions, changes the protein specificity to bindthe UTR, stabilizes against nucleases, and/or improves cellulartolerance (e.g., leads to a reduced innate immune response). In otherembodiments, addition and/or swapping of the RNA binding module of theR2 protein is compatible with the use of different sequence or ligandsthat are linked to the transgene and/or element module of the RNA. Insome embodiments, combinations of new ligands in place of the UTRs willhave better affinity to the RNA binding domain of R2 and lead to betterinsertion efficiency. In some embodiments, the changes to the sequenceof the UTRs or changes to the base modifications of the UTRs willincrease the secondary structure stability that leads to betterinteraction with the RNA binding module.

Example 22: Assays to Measure RNA Binding Affinity to New Sequences

New UTR modules are tested in a binding assay. In the case of new RNAs,they are synthesized either by cell-free in vitro transcription using asynthetic DNA template or by chemical synthesis of the RNA infull-length or chemical synthesis of pieces that are ligated together toform a single RNA molecule. The binding affinity of the purified UTRsare measured in a binding assay (e.g., EMSA, Fluorescence anisotropy,dual-filter binding, FRET, SPR, or thermophoresis (temperature relatedintensity change)). The UTR module and/or RNA binding module/RT moduleis detected with or without a label which is described above forlabeling RNAs. Measurement of the molecules at different concentrationsis performed to determine a binding affinity. In some embodiments,alterations to/swapping of the 5′ and/or 3′ UTR binding module and/orchanges to the RNA binding/RT module will lead to better interactionsthan the wild-type R2 protein or UTR. In some embodiments, the increasedinteraction will lead to an increase in the efficiency ofretro-transposition and in some cases increases specificity of the R2protein to interact with the RNA.

Example 23: Alternative UTRs

While not wishing to be bound by theory, in some embodiments the UTRsact as a handle for the R2 protein to interact with the RNA which ituses as a template for RT in concert with it binding a genomic location,nicking the DNA with its endonuclease module, and then using the boundRNA as a template for RT insertion at the cleavage site in the DNA. Forthe UTR to keep the template in close proximity to the RT module, thenthe UTR modules can be substituted with different ligands that wouldbind to a specific RNA binding module engineered into the R2 protein.Thus, in some embodiments, the alternative non-RNA UTR is either aprotein, small molecule, or other chemical entity that is attachedcovalently, through protein-protein interaction, small molecule-proteininteraction, or through hybridization. In some embodiments the RNAbinding module binds specifically to a ligand that is not RNA that isattached to the transgene module RNA that increases the efficiency,stability, and/or rate of retro-transposition.

Example 24: Assays to Measure the Activity of UTR Constructs

Binding assays to measure affinity of R2 protein with engineered UTRsare performed as described above, e.g., for a protein-nucleic acidinteraction. In cases of protein-protein or protein-small moleculeinteractions the assay uses a label on the RNA transgene module wherethe UTR module is attached.

Example 25: Targeted Genomic Integration

In this example, Gene Writing technology is delivered to target cellsand to non-target cells, and new DNA is integrated into the genome intarget cells at a higher frequency than in non-target cells. Asdescribed in more detail below, this approach takes advantage of thenon-target cell having an endogenous miRNA that the target cell does nothave (or has at a lower level). The endogenous miRNA is used to reduceDNA integration in the non-target cell.

The polypeptide used is the R2Tg protein and the template RNA componentis RNA coding for the GFP protein and flanked at the 5′ end by the 5′UTR and at the 3′ end by the 3′ UTR of the R2Tg retrotransposase. The 5′UTR is flanked by 100 bp of homology to the 5′ of R2Tg 28s rDNA targetsite and the 3′ UTR is flanked by 100 bp of homology to the 3′ of R2Tg28s rDNA target site. The GFP gene is facing in the antisense directionwith regard to the 5′ and 3′ UTRs and has its own promoter andpolyadenylation signal.

The template RNA further comprises a microRNA recognition sequence. ThismicroRNA recognition sequence is bound by microRNAs in the non-targetcells, leading to the inhibition (e.g., degradation) of the template RNAprior to genomic integration.

In this example the target cells are hepatocytes and the non-targetcells are macrophages from the hematopoietic lineage. The target cellsand non-target cells are cultured separately. The template RNA andretrotransposase protein can be delivered to cells as described herein,e.g., as RNA or using viral vectors (e.g. adeno-associated viralvectors), wherein the template RNA is transcribed from viral vector DNA.

Three days after treating the cells, GFP expression and genomicintegration are assayed.

GFP expression is assayed via flow cytometry. In some embodiments, GFPexpression will be higher in the hepatocyte population than in themacrophage population.

Genomic integration (in terms of copy number per cell normalized to areference gene) is assayed via droplet digital PCR using methodsdescribed herein. In some embodiments, genomic integration will behigher in the hepatocyte population than in the macrophage population.

Example 26: Testing Modularity of the DNA Binding Domain

In this example, a series of experiments were performed to test theactivity of various mutant retrotransposases, as well as gainingstructural knowledge about these proteins. This experiments testedflexible linkers in different locations and lengths, in order todetermine if the DNA binding domain (DBD) was modular. These experimentsalso provide support for being able to separate the DBD from the rest ofR2Tg and replacing it with any DNA targeting protein sequence. Thisexample thus supports an understanding that the transposases describedherein can withstand the tested levels of sequence divergence at aplurality of locations (e.g., in the predicted −1 RNA binding motif, inan alpha helix, and in a coil region located C-terminal to the predictedc-myb DNA binding motif, e.g., as described below) identified bystructural modeling, while maintaining function.

Briefly, the two linkers (Linker A: SGSETPGTSESATPES (SEQ ID NO: 1023),and Linker B: GGGS (SEQ ID NO: 1024)) were inserted into 3 locations,noted herein as versions v1, v2, and v3. v1 was located at theN-terminal side of an alpha helical region of R2Tg that preceded thepredicted −1 RNA binding motif, v2 was located at the C-terminal side ofan alpha helical region of R2Tg that preceded the predicted −1 RNAbinding motif, and v3 was located C-terminal to a random coil regionthat came after the predicted c-myb DNA binding motif of R2Tg. For eachof v1, v2, and v3, one of linkers A or B were added by PCR to a DNAplasmid that expressed R2Tg, thereby yielding sequences v1A (v1+linkerA), v1B (v1+linker B), v1C (v1+linker C), v2A (v2+linker A), v2B(v2+linker B), and v2C (v2+linker C), as shown in Table 5 below. Theinsertion of the linkers was verified by Sanger sequencing and the DNAplasmids were purified for transfection.

TABLE 5 Amino acid sequences of R2Tg mutants with linkers in the DNAbinding domain (DBD) R2Tg SEQ Mutant + ID Linker Amino Acid Sequence NOR2Tg MASCPKPGPPVSAGAMSLESGLTTHSVLAIERGPNS 1017 with DBDLANSGSDFGGGGLGLPLRLLRVSVGTQTSRSDWVDL LinkerVSWSHPGPTSKSQQVDLVSLFPKHRVDLLSKNDQVD v1ALVAQFLPSKFPPNLAENDLALLVNLEFYRSDLHVYECVHFAAHWEGLSGLPEVYEQLAPQPCVGETLHSSLPRDSELFVPEEGSSEKESEDAPKTSPPTPGKHGLEQTGEEKVMVTVPDKNPPCPCCGTRVNSVLNLIEHLKVSHGKRGVCFRCAKCGKENSNYHSVVCHFPKCRGPETEKAPAGEWICEVCNRDFTTKIGLGQHKRLAHPAVRNQERIVASQPKETSNRGAHKRCWTKEEEELLIRLEAQFEGNKNINKLIAEHITTKTAKQISDKRRLLSRKPAEEPREEPGTCHHTRRAAASLRTEPEMSHHAQAEDRDNGPGRRPLPGRAAAGGRTMDEIRRHPDKGNGQQRPTKQKSEEQLQAYYKKTLEERLSAGALNTFPRAFKQVMEGRDIKLVINQTAQDSGSETPGTSESATPESCFGCLESISQIRTATRDKKDTVTREKHPKKPFQKWMKDRAIKKGNYLRFQRLFYLDRGKLAKIILDDIECLSCDIPLSEIYSVFKTRWETTGSFKSLGDFKTYGKADNTAFRELITAKEIEKNVQEMSKGSAPGPDGITLGDVVKMDPEFSRTMEIFNLWLTTGKIPDMVRGCRTVLIPKSSKPDRLKDINNWRPITIGSILLRLFSRIVTARLSKACPLNPRQRGFIRAAGCSENLKLLQTIIWSAKREHRPLGVVFVDIAKAFDTVSHQHIIHALQQREVDPHIVGLVSNMYENISTYITTKRNTHTDKIQIRVGVKQGDPMSPLLFNLAMDPLLCKLEESGKGYHRGQSSITAMAFADDLVLLSDSWENMNTNISILETFCNLTGLKTQGQKCHGFYIKPTKDSYTINDCAAWTINGTPLNMIDPGESEKYLGLQFDPWIGIARSGLSTKLDFWLQRIDQAPLKPLQKTDILKTYTIPRLIYIADHSEVKTALLETLDQKIRTAVKEWLHLPPCTCDAILYSSTRDGGLGITKLAGLIPSVQARRLHRIAQSSDDTMKCFMEKEKMEQLHKKLWIQAGGDRENIPSIWEAPPSSEPPNNVSTNSEWEAPTQKDKFPKPCNWRKNEFKKWTKLASQGRGIVNFERDKISNHWIQYYRRIPHRKLLTALQLRANVYPTREFLARGRQDQYIKACRHCDADIESCAHIIGNCPVTQDARIKRHNYICELLLEEAKKKDWVVFKEPHIRDSNKELYKPDLIFVKDARALVVDVTVRYEAAKSSLEEAAAEKVRKYKHLETEVRHLTNAKDVTFVGFPLGARGKWHQDNFKLLTELGLSKSRQVKMAETFSTVALFSSVDIVHMFASRARKSM VM R2TgMASCPKPGPPVSAGAMSLESGLTTHSVLAIERGPNS 1018 with DBDLANSGSDFGGGGLGLPLRLLRVSVGTQTSRSDWVDL LinkerVSWSHPGPTSKSQQVDLVSLFPKHRVDLLSKNDQVD v1BLVAQFLPSKFPPNLAENDLALLVNLEFYRSDLHVYECVHFAAHWEGLSGLPEVYEQLAPQPCVGETLHSSLPRDSELFVPEEGSSEKESEDAPKTSPPTPGKHGLEQTGEEKVMVTVPDKNPPCPCCGTRVNSVLNLIEHLKVSHGKRGVCFRCAKCGKENSNYHSVVCHFPKCRGPETEKAPAGEWICEVCNRDFTTKIGLGQHKRLAHPAVRNQERIVASQPKETSNRGAHKRCWTKEEEELLIRLEAQFEGNKNINKLIAEHITTKTAKQISDKRRLLSRKPAEEPREEPGTCHHTRRAAASLRTEPEMSHHAQAEDRDNGPGRRPLPGRAAAGGRTMDEIRRHPDKGNGQQRPTKQKSEEQLQAYYKKTLEERLSAGALNTFPRAFKQVMEGRDIKLVINQTAQDGGGSCFGCLESISQIRTATRDKKDTVTREKHPKKPFQKWMKDRAIKKGNYLRFQRLFYLDRGKLAKIILDDIECLSCDIPLSEIYSVFKTRWETTGSFKSLGDFKTYGKADNTAFRELITAKEIEKNVQEMSKGSAPGPDGITLGDVVKMDPEFSRTMEIFNLWLTTGKIPDMVRGCRTVLIPKSSKPDRLKDINNWRPITIGSILLRLFSRIVTARLSKACPLNPRQRGFIRAAGCSENLKLLQTIIWSAKREHRPLGVVFVDIAKAFDTVSHQHIIHALQQREVDPHIVGLVSNMYENISTYITTKRNTHTDKIQIRVGVKQGDPMSPLLFNLAMDPLLCKLEESGKGYHRGQSSITAMAFADDLVLLSDSWENMNTNISILETFCNLTGLKTQGQKCHGFYIKPTKDSYTINDCAAWTINGTPLNMIDPGESEKYLGLQFDPWIGIARSGLSTKLDFWLQRIDQAPLKPLQKTDILKTYTIPRLIYIADHSEVKTALLETLDQKIRTAVKEWLHLPPCTCDAILYSSTRDGGLGITKLAGLIPSVQARRLHRIAQSSDDTMKCFMEKEKMEQLHKKLWIQAGGDRENIPSIWEAPPSSEPPNNVSTNSEWEAPTQKDKFPKPCNWRKNEFKKWTKLASQGRGIVNFERDKISNHWIQYYRRIPHRKLLTALQLRANVYPTREFLARGRQDQYIKACRHCDADIESCAHIIGNCPVTQDARIKRHNYICELLLEEAKKKDWVVFKEPHIRDSNKELYKPDLIFVKDARALVVDVTVRYEAAKSSLEEAAAEKVRKYKHLETEVRHLTNAKDVTFVGFPLGARGKWHQDNFKLLTELGLSKSRQVKMAET FSTVALFSSVDIVHMFASRARKSMVM R2TgMASCPKPGPPVSAGAMSLESGLTTHSVLAIERGPNS 1019 with DBDLANSGSDFGGGGLGLPLRLLRVSVGTQTSRSDWVDL LinkerVSWSHPGPTSKSQQVDLVSLFPKHRVDLLSKNDQVD v2ALVAQFLPSKFPPNLAENDLALLVNLEFYRSDLHVYECVHFAAHWEGLSGLPEVYEQLAPQPCVGETLHSSLPRDSELFVPEEGSSEKESEDAPKTSPPTPGKHGLEQTGEEKVMVTVPDKNPPCPCCGTRVNSVLNLIEHLKVSHGKRGVCFRCAKCGKENSNYHSVVCHFPKCRGPETEKAPAGEWICEVCNRDFTTKIGLGQHKRLAHPAVRNQERIVASQPKETSNRGAHKRCWTKEEEELLIRLEAQFEGNKNINKLIAEHITTKTAKQISDKRRLLSRKPAEEPREEPGTCHHTRRAAASLRTEPEMSHHAQAEDRDNGPGRRPLPGRAAAGGRTMDEIRRHPDKGNGQQRPTKQKSEEQLQAYYKKTLEERLSAGALNTFPRAFKQVMEGRDIKLVINQTAQDCFGCLESISQIRSGSETPGTSESATPESTATRDKKDTVTREKHPKKPFQKWMKDRAIKKGNYLRFQRLFYLDRGKLAKIILDDIECLSCDIPLSEIYSVFKTRWETTGSFKSLGDFKTYGKADNTAFRELITAKEIEKNVQEMSKGSAPGPDGITLGDVVKMDPEFSRTMEIFNLWLTTGKIPDMVRGCRTVLIPKSSKPDRLKDINNWRPITIGSILLRLFSRIVTARLSKACPLNPRQRGFIRAAGCSENLKLLQTIIWSAKREHRPLGVVFVDIAKAFDTVSHQHIIHALQQREVDPHIVGLVSNMYENISTYITTKRNTHTDKIQIRVGVKQGDPMSPLLFNLAMDPLLCKLEESGKGYHRGQSSITAMAFADDLVLLSDSWENMNTNISILETFCNLTGLKTQGQKCHGFYIKPTKDSYTINDCAAWTINGTPLNMIDPGESEKYLGLQFDPWIGIARSGLSTKLDFWLQRIDQAPLKPLQKTDILKTYTIPRLIYIADHSEVKTALLETLDQKIRTAVKEWLHLPPCTCDAILYSSTRDGGLGITKLAGLIPSVQARRLHRIAQSSDDTMKCFMEKEKMEQLHKKLWIQAGGDRENIPSIWEAPPSSEPPNNVSTNSEWEAPTQKDKFPKPCNWRKNEFKKWTKLASQGRGIVNFERDKISNHWIQYYRRIPHRKLLTALQLRANVYPTREFLARGRQDQYIKACRHCDADIESCAHIIGNCPVTQDARIKRHNYICELLLEEAKKKDWVVFKEPHIRDSNKELYKPDLIFVKDARALVVDVTVRYEAAKSSLEEAAAEKVRKYKHLETEVRHLTNAKDVTFVGFPLGARGKWHQDNFKLLTELGLSKSRQVKMAETFSTVALFSSVDIVHMFASRARKSM VM R2TgMASCPKPGPPVSAGAMSLESGLTTHSVLAIERGPNS 1020 with DBDLANSGSDFGGGGLGLPLRLLRVSVGTQTSRSDWVDL LinkerVSWSHPGPTSKSQQVDLVSLFPKHRVDLLSKNDQVD v2BLVAQFLPSKFPPNLAENDLALLVNLEFYRSDLHVYECVHFAAHWEGLSGLPEVYEQLAPQPCVGETLHSSLPRDSELFVPEEGSSEKESEDAPKTSPPTPGKHGLEQTGEEKVMVTVPDKNPPCPCCGTRVNSVLNLIEHLKVSHGKRGVCFRCAKCGKENSNYHSVVCHFPKCRGPETEKAPAGEWICEVCNRDFTTKIGLGQHKRLAHPAVRNQERIVASQPKETSNRGAHKRCWTKEEEELLIRLEAQFEGNKNINKLIAEHITTKTAKQISDKRRLLSRKPAEEPREEPGTCHHTRRAAASLRTEPEMSHHAQAEDRDNGPGRRPLPGRAAAGGRTMDEIRRHPDKGNGQQRPTKQKSEEQLQAYYKKTLEERLSAGALNTFPRAFKQVMEGRDIKLVINQTAQDCFGCLESISQIRGGGSTATRDKKDTVTREKHPKKPFQKWMKDRAIKKGNYLRFQRLFYLDRGKLAKIILDDIECLSCDIPLSEIYSVFKTRWETTGSFKSLGDFKTYGKADNTAFRELITAKEIEKNVQEMSKGSAPGPDGITLGDVVKMDPEFSRTMEIFNLWLTTGKIPDMVRGCRTVLIPKSSKPDRLKDINNWRPITIGSILLRLFSRIVTARLSKACPLNPRQRGFIRAAGCSENLKLLQTIIWSAKREHRPLGVVFVDIAKAFDTVSHQHIIHALQQREVDPHIVGLVSNMYENISTYITTKRNTHTDKIQIRVGVKQGDPMSPLLFNLAMDPLLCKLEESGKGYHRGQSSITAMAFADDLVLLSDSWENMNTNISILETFCNLTGLKTQGQKCHGFYIKPTKDSYTINDCAAWTINGTPLNMIDPGESEKYLGLQFDPWIGIARSGLSTKLDFWLQRIDQAPLKPLQKTDILKTYTIPRLIYIADHSEVKTALLETLDQKIRTAVKEWLHLPPCTCDAILYSSTRDGGLGITKLAGLIPSVQARRLHRIAQSSDDTMKCFMEKEKMEQLHKKLWIQAGGDRENIPSIWEAPPSSEPPNNVSTNSEWEAPTQKDKFPKPCNWRKNEFKKWTKLASQGRGIVNFERDKISNHWIQYYRRIPHRKLLTALQLRANVYPTREFLARGRQDQYIKACRHCDADIESCAHIIGNCPVTQDARIKRHNYICELLLEEAKKKDWVVFKEPHIRDSNKELYKPDLIFVKDARALVVDVTVRYEAAKSSLEEAAAEKVRKYKHLETEVRHLTNAKDVTFVGFPLGARGKWHQDNFKLLTELGLSKSRQVKMAET FSTVALFSSVDIVHMFASRARKSMVM R2TgMASCPKPGPPVSAGAMSLESGLTTHSVLAIERGPNS 1021 with DBDLANSGSDFGGGGLGLPLRLLRVSVGTQTSRSDWVDL LinkerVSWSHPGPTSKSQQVDLVSLFPKHRVDLLSKNDQVD v3ALVAQFLPSKFPPNLAENDLALLVNLEFYRSDLHVYECVHFAAHWEGLSGLPEVYEQLAPQPCVGETLHSSLPRDSELFVPEEGSSEKESEDAPKTSPPTPGKHGLEQTGEEKVMVTVPDKNPPCPCCGTRVNSVLNLIEHLKVSHGKRGVCFRCAKCGKENSNYHSVVCHFPKCRGPETEKAPAGEWICEVCNRDFTTKIGLGQHKRLAHPAVRNQERIVASQPKETSNRGAHKRCWTKEEEELLIRLEAQFEGNKNINKLIAEHITTKTAKQISDKRRLLSRKPAEEPREEPGTCHHTRRAASGSETPGTSESATPESASLRTEPEMSHHAQAEDRDNGPGRRPLPGRAAAGGRTMDEIRRHPDKGNGQQRPTKQKSEEQLQAYYKKTLEERLSAGALNTFPRAFKQVMEGRDIKLVINQTAQDCFGCLESISQIRTATRDKKDTVTREKHPKKPFQKWMKDRAIKKGNYLRFQRLFYLDRGKLAKIILDDIECLSCDIPLSEIYSVFKTRWETTGSFKSLGDFKTYGKADNTAFRELITAKEIEKNVQEMSKGSAPGPDGITLGDVVKMDPEFSRTMEIFNLWLTTGKIPDMVRGCRTVLIPKSSKPDRLKDINNWRPITIGSILLRLFSRIVTARLSKACPLNPRQRGFIRAAGCSENLKLLQTIIWSAKREHRPLGVVFVDIAKAFDTVSHQHIIHALQQREVDPHIVGLVSNMYENISTYITTKRNTHTDKIQIRVGVKQGDPMSPLLFNLAMDPLLCKLEESGKGYHRGQSSITAMAFADDLVLLSDSWENMNTNISILETFCNLTGLKTQGQKCHGFYIKPTKDSYTINDCAAWTINGTPLNMIDPGESEKYLGLQFDPWIGIARSGLSTKLDFWLQRIDQAPLKPLQKTDILKTYTIPRLIYIADHSEVKTALLETLDQKIRTAVKEWLHLPPCTCDAILYSSTRDGGLGITKLAGLIPSVQARRLHRIAQSSDDTMKCFMEKEKMEQLHKKLWIQAGGDRENIPSIWEAPPSSEPPNNVSTNSEWEAPTQKDKFPKPCNWRKNEFKKWTKLASQGRGIVNFERDKISNHWIQYYRRIPHRKLLTALQLRANVYPTREFLARGRQDQYIKACRHCDADIESCAHIIGNCPVTQDARIKRHNYICELLLEEAKKKDWVVFKEPHIRDSNKELYKPDLIFVKDARALVVDVTVRYEAAKSSLEEAAAEKVRKYKHLETEVRHLTNAKDVTFVGFPLGARGKWHQDNFKLLTELGLSKSRQVKMAETFSTVALFSSVDIVHMFASRARKSM VM R2TgMASCPKPGPPVSAGAMSLESGLTTHSVLAIERGPNS 1022 with DBDLANSGSDFGGGGLGLPLRLLRVSVGTQTSRSDWVDL LinkerVSWSHPGPTSKSQQVDLVSLFPKHRVDLLSKNDQVD v3BLVAQFLPSKFPPNLAENDLALLVNLEFYRSDLHVYECVHFAAHWEGLSGLPEVYEQLAPQPCVGETLHSSLPRDSELFVPEEGSSEKESEDAPKTSPPTPGKHGLEQTGEEKVMVTVPDKNPPCPCCGTRVNSVLNLIEHLKVSHGKRGVCFRCAKCGKENSNYHSVVCHFPKCRGPETEKAPAGEWICEVCNRDFTTKIGLGQHKRLAHPAVRNQERIVASQPKETSNRGAHKRCWTKEEEELLIRLEAQFEGNKNINKLIAEHITTKTAKQISDKRRLLSRKPAEEPREEPGTCHHTRRAAGGGSASLRTEPEMSHHAQAEDRDNGPGRRPLPGRAAAGGRTMDEIRRHPDKGNGQQRPTKQKSEEQLQAYYKKTLEERLSAGALNTFPRAFKQVMEGRDIKLVINQTAQDCFGCLESISQIRTATRDKKDTVTREKHPKKPFQKWMKDRAIKKGNYLRFQRLFYLDRGKLAKIILDDIECLSCDIPLSEIYSVFKTRWETTGSFKSLGDFKTYGKADNTAFRELITAKEIEKNVQEMSKGSAPGPDGITLGDVVKMDPEFSRTMEIFNLWLTTGKIPDMVRGCRTVLIPKSSKPDRLKDINNWRPITIGSILLRLFSRIVTARLSKACPLNPRQRGFIRAAGCSENLKLLQTIIWSAKREHRPLGVVFVDIAKAFDTVSHQHIIHALQQREVDPHIVGLVSNMYENISTYITTKRNTHTDKIQIRVGVKQGDPMSPLLFNLAMDPLLCKLEESGKGYHRGQSSITAMAFADDLVLLSDSWENMNTNISILETFCNLTGLKTQGQKCHGFYIKPTKDSYTINDCAAWTINGTPLNMIDPGESEKYLGLQFDPWIGIARSGLSTKLDFWLQRIDQAPLKPLQKTDILKTYTIPRLIYIADHSEVKTALLETLDQKIRTAVKEWLHLPPCTCDAILYSSTRDGGLGITKLAGLIPSVQARRLHRIAQSSDDTMKCFMEKEKMEQLHKKLWIQAGGDRENIPSIWEAPPSSEPPNNVSTNSEWEAPTQKDKFPKPCNWRKNEFKKWTKLASQGRGIVNFERDKISNHWIQYYRRIPHRKLLTALQLRANVYPTREFLARGRQDQYIKACRHCDADIESCAHIIGNCPVTQDARIKRHNYICELLLEEAKKKDWVVFKEPHIRDSNKELYKPDLIFVKDARALVVDVTVRYEAAKSSLEEAAAEKVRKYKHLETEVRHLTNAKDVTFVGFPLGARGKWHQDNFKLLTELGLSKSRQVKMAET FSTVALFSSVDIVHMFASRARKSMVM

HEK293T cells were plated in 96-well plates and grown overnight at 37°C., 5% CO2. The HEK293T cells were transfected with plasmids thatexpressed R2Tg (wild-type), R2 endonuclease mutant, and linker mutants.The transfection was carried out using the Fugene HD transfectionreagent according to the manufacturer recommendations, where each wellreceived 80 ng of plasmid DNA and 0.5 μL of transfection reagent. Alltransfections were performed in duplicate and the cells were incubatedfor 72 h prior to genomic DNA extraction.

Activity of the mutants was measured by a ddPCR assay that quantifiedthe copy number of R2Tg integration per genome. The 5′ and 3′ junctionswere quantified by generating two different amplicons at each end.

v3 (near the c-myb binding motif in the DBD) decreased integrationactivity with either linker A or B. v1 (N-terminal to the alpha helixpreceding the −1 RNA binding motif) had comparable activity to thewild-type when used with linker A (16 AA) versus the shorter linker B (4AA). This could be related to amino acid selection, length, orthree-dimensional structure. v2 (C-terminal to the alpha helix precedingthe −1 RNA binding motif) did not tolerate linker A; however, linker Bhad activity that was comparable and slightly better than the wild-type.v1 and v2 may therefore be considered preferred locations to add alinker that can separate R2Tg's DNA binding domain and the rest of theprotein.

Example 27: Long-Read Sequencing to Determine Integration Fidelity

Retrotransposon integration experiments were performed as described inprevious examples. In one example, PCR amplification was used togenerate amplicons by designing one primer targeting the genomicintegration site and one primer targeting the integrant sequence. Inthis example, these primers were designed to maximize the length of theamplified genomic locus fused with the integrant sequence. By poolingamplicons spanning both ends of the integrant and performing long-readnext-generation sequencing, the fidelity of each integration wasevaluated.

A cis construct of R2Tg was integrated into 293T cells via plasmidtransfection as described herein. Amplicons spanning each end of theintegrations were generated with flanking randomized UMIs to control forPCR bias. These amplicons were sequenced with PacBio next-generationsequencing. The resulting sequences were collapsed to remove reads withidentical UMIs. By aligning unique reads, a coverage plot wasconstructed as shown in FIGS. 20A-20B. Sequence coverage largely showsuniform coverage across amplicons, indicating significant fidelity ofintegration. An associated reverse-transcriptase deficient mutantcontrol produced no signal. Internal deletions were also analyzed inFIGS. 21A-21B. Internal deletions were generally low relative to overallunique read counts, with some clustering at the 5′ junction ofrDNA-R2Tg.

In another example, hybrid capture may be performed as described in aprevious example but with a larger target library length during initiallibrary generation. The resulting library can then be subjected tolong-read next-generation sequencing.

Example 28: Targeted Delivery of R2Gfo and R4A1 Retrotransposon toMammalian Cells

This example describes targeted integration of the R2Gfo and R4A1retrotransposon elements to mammalian cells via DNA delivery.

In one example, we assayed the full R2 element R2-1_GFo (Repbase; Kojimaet al PLoS One 11, e0163496 (2015)) from the medium ground finch,Geospiza fortis (“R2GFo”). In another example, we assayed the full R4element R4_AL (Repbase; Burke et al Nucleic Acids Res. 23, 4628-34(1995)) from the large roundworm, Ascaris lumbricoides (“R4A1”). Becausenon-LTR R2 and R4 elements are not present in the human genome and arethought to be highly site-specific, the ability of retrotransposons toaccurately and efficiently integrate itself into the human genome woulddemonstrate the capability to perform genomic targeted integration.

Plasmids harboring R2Gfo (PLV033) or R4A1 (PLV462) were designed for cisintegration of the R2Gfo or R4A1 elements as in previous examples.Plasmids were synthesized such that the wildtype element was flanked byits native un-translated regions (UTRs) and 100 bp of homology to itsrDNA target (FIG. 22). The element expression was driven by themammalian CMV promoter. We introduced each plasmid into HEK393T cellsusing the FuGENE® HD transfection reagent. HEK293T cells were seeded in96-well plates at 10,000 cells/well 24 hours before transfection. On thetransfection day, 0.50 transfection reagent and 80 ng DNA was mixed in10 μl Opti-MEM and incubated for 15 minutes at room temperature. Thetransfection mixture was then added to the medium of the seeded cells.Three days after transfection, genomic DNA was extracted forretrotransposition assays. R2Tg was also delivered in parallel in thesame format to serve as a comparison.

ddPCR was performed to confirm integration and assess integrationefficiency. A Taqman probe was designed to the 3′UTR portion of eachelement. A forward primer was synthesized to bind directly upstream ofthe probe, and a reverse primer was synthesized to bind the rDNA. Thus,amplification of the expected product across the integration junctionwould degrade the probe and create a fluorescent signal. The results ofthe ddPCR copy number analysis (in comparison to reference gene RPP30)are shown in FIG. 23. R2Gfo integration achieved a mean copy number of0.21 integrants/genome in this experiment. R4A1 achieved a mean copynumber of 0.085 integrants/genome.

Example 29: Integration of Retrotransposons into Human Fibroblasts

This example describes the cis integration of R2Tg into humanfibroblasts. Briefly, a plasmid designed to integrate R2Tg in cis wassynthesized such that R2Tg was flanked by its native UTRs and homologoussequence to its rDNA target as in previous examples. 0.5 μg PLV014(wild-type) and PLV072 (EN mutant) plasmids were transfected into100,000 human dermal fibroblasts isolated from neonatal foreskin (HDFn,C0045C, ThermoFisher Scientific) respectively using the Neontransfection system. Two programs were performed, each in duplicate. Thesetting for Program 1 was 1700V pulse voltage, 20 ms pulse width, and 1pulse number. The setting for Program 2 was 1400V pulse voltage, 20 mspulse width, and 2 pulse number. Both programs achieved 95% transfectionefficiency measured using plasmid encoding the EGFP. Three days posttransfection, genomic DNA was extracted for the ddPCR assay. ddPCR wasperformed to confirm integration and assess integration efficiency. ATaqman probe was designed to the 3′ UTR portion of the R2Tg element. Aforward primer was synthesized to bind directly upstream of the probe,and a reverse primer was synthesized to bind the rDNA. Thus,amplification of the expected product across the integration junctionwould degrade the probe and create a fluorescent signal. The results ofddPCR copy number analysis (in comparison to reference gene RPP30) areshown in FIG. 24. Wild-type (WT) R2Tg integration achieved a mean copynumber of 0.036 integrants/genome in this experiment, significantlyhigher than a control R2Tg plasmid with a point mutation abolishingendonuclease activity (EN).

Example 30: Evaluation of DNA Damage Response Upon RetrotransposonTransfection

DNA damage (e.g., resulting from DSB formation or replication forkcollapse) leads to the activation of p53, which among many othertranscriptional responses, leads to the upregulation of p21, resultingin cell cycle arrest or apoptosis. Genome editing using CRISRP/Cas9 hasbeen shown to activate p53 and p21, which is a potential safety andefficacy problem for CRISPR/based therapeutics. To establish whetherR2Tg delivery to the cell leads to activation of p53 and p21, U2OS cellswere seeded at a density of 4×10⁴ cells/well and transfected 24 hourslater using the Fugene HD and Lipofectamine reagents with either 500 ngof R2Tg-WT plasmid or 500 ng of R2Tg-EN (a variant of R2Tg with amutation in the endonuclease (EN) domain, rendering R2Tg inactive). Tocontrol for transfection efficiency, U2OS cells were also transfectedwith a plasmid expressing GFP. Lastly, as a positive control for p53 andp21 activation, U2OS cells were treated with one of the DNAdamage-inducing agents etoposide (20 μM) or bleomycin (10 μg/ml). TheU2OS cells were collected 24 hours after transfection/treatment. Proteinlysates were prepared in RIPA buffer and run on an SDS-PAGE gel,followed by transfer to nitrocellulose, followed by probing withantibodies against p53 and p21, as well as Actin and Vinculin. As shownin FIG. 25, no R2Tg-induced upregulation of p53 or p21 above the GFPplasmid control was detected in either transfection condition.

LENGTHY TABLES The patent application contains a lengthy table section.A copy of the table is available in electronic form from the USPTO website(http://seqdata.uspto.gov/?pageRequest=docDetail&DocID=US20200109398A1).An electronic copy of the table will also be available from the USPTOupon request and payment of the fee set forth in 37 CFR 1.19(b)(3).

What is claimed is:
 1. A system for modifying DNA comprising: apolypeptide or a nucleic acid encoding a polypeptide capable of targetprimed reverse transcription, wherein the polypeptide comprises (a) areverse transcriptase domain and (b) an endonuclease domain, wherein atleast one of (a) or (b) is heterologous, and a template RNA comprising(i) a sequence that binds the polypeptide and (ii) a heterologous objectsequence.
 2. A system for modifying DNA comprising: a polypeptide or anucleic acid encoding a polypeptide capable of target primed reversetranscription, wherein the polypeptide comprises (a) a target DNAbinding domain, (b) a reverse transcriptase domain and (c) anendonuclease domain, wherein at least one of (a), (b) or (c) isheterologous, and a template RNA comprising (i) a sequence that bindsthe polypeptide and (ii) a heterologous object sequence.
 3. The systemof claim 1, wherein the polypeptide comprises a sequence of at least 50amino acids having at least 80% identity to a reverse transcriptasedomain of a sequence of a polypeptide listed in TABLE 1, TABLE 2, orTABLE
 3. 4. The system of claim 1, wherein the reverse transcriptasedomain is from a retrovirus or a retrotransposon, such as aLTR-retrotransposon, or a non-LTR retrotransposon.
 5. The system ofclaim 4, wherein the reverse transcriptase is from a non-LTRretrotransposon, wherein the non-LTR retrotransposon is: a RLE-typenon-LTR retrotransposon from the R2, NeSL, HERO, R4, or CRE clade, or anAPE-type non-LTR retrotransposon from the R1, or Tx1 clade.
 6. Thesystem of claim 1, wherein the reverse transcriptase domain is from anavian retrotransposase of column 8 of Table 3, or a sequence having atleast 70%, identity thereto.
 7. The system of claim 1, wherein theendonuclease domain is heterologous to the reverse transcriptase domain,and wherein the endonuclease is a Fok1 nuclease (or a functionalfragment thereof), a type-II restriction 1-like endonuclease (RLE-typenuclease), another RLE-type endonuclease, or a Prp8 nuclease.
 8. Thesystem of claim 1, wherein the endonuclease domain is heterologous tothe reverse transcriptase domain, wherein endonuclease domain containsDNA binding functionality.
 9. The system of claim 1, wherein theendonuclease domain is heterologous to the reverse transcriptase domain,and wherein the endonuclease has nickase activity and does not formdouble stranded breaks.
 10. The system of claim 1, wherein thepolypeptide comprises a DNA binding domain heterologous to the reversetranscriptase domain, and wherein the DNA binding domain is: azinc-finger element, or a functional fragment thereof; or a TAL effectorelement, or a functional fragment thereof; a Myb domain; or asequence-guided DNA binding element.
 11. The system of claim 1, whereinthe polypeptide comprises a DNA binding domain heterologous to thereverse transcriptase domain, and wherein the DNA binding element is asequence-guided DNA binding element, further wherein the sequence-guidedDNA binding element is Cas9, Cpf1, or other CRISPR-related protein. 12.The system of claim 11, wherein the sequence-guided DNA binding elementhas been altered to have no endonuclease activity.
 13. The system ofclaim 11, wherein the sequence-guided DNA binding element replaces theendonuclease element of the polypeptide.
 14. The system of any of claim1, wherein the reverse transcriptase domain does not comprise an RNAbinding domain and the polypeptide comprises an RNA binding domainheterologous to the reverse transcriptase domain, wherein the RNAbinding domain is a B-box protein, a MS2 coat protein, a dCas protein,or a UTR binding protein, or a fragment or variant of any of theforegoing.
 15. The system of claim 1, wherein the polypeptide comprisesa DNA binding domain heterologous to the reverse transcriptase domain,and wherein the DNA binding domain is a transcription factor.
 16. Thesystem of any of claim 1, which is capable of modifying DNA usingreverse transcriptase activity, optionally in the absence of homologousrecombination activity.
 17. The system of any of the foregoing claims,which is capable of modifying DNA by insertion of the heterologousobject sequence without an intervening DNA-dependent RNA polymerizationof the template RNA.
 18. The system of claim 1, wherein the domains ofthe polypeptide are joined by a linker; or wherein the domains of thepolypeptide are joined by a peptide linker; or wherein the domains ofthe polypeptide are joined by a peptide linker and the peptide linker isat least 2 amino acids in length; or wherein the DNA binding domain isattached to the reverse transcriptase domain via a synthetic linker; orwherein the endonuclease domain attached to the reverse transcriptiondomain via a synthetic linker.
 19. The system of claim 1, wherein thepolypeptide further comprises a nuclear localization sequence.
 20. Thesystem of any of claim 10, wherein the reverse transcriptase domain,endonuclease domain, or DNA binding domain are modified; or the reversetranscriptase domain is mutated to bind a heterologous template RNA; orthe endonuclease domain is mutated to alter DNA endonuclease activity;or the DNA binding domain is modified to alter DNA-binding specificityand affinity.
 21. The system of any of the preceding claims, wherein thetemplate RNA comprises one or more of a microRNA sequence, a siRNAsequence, a guide RNA sequence, a piwi RNA sequence; or wherein thetemplate RNA comprises a guide RNA sequence.
 22. A method of modifying atarget DNA strand in a cell, tissue or subject, comprising administeringthe system of claim 1 to the cell, tissue or subject, wherein the systemreverse transcribes the template RNA sequence into the target DNAstrand, thereby modifying the target DNA strand.
 23. The method of claim22, wherein the sequence that binds the polypeptide has one or more ofthe following characteristics: (a) is at the 3′ end of the template RNA;(b) is a non-coding sequence; (c) is a structured RNA; (d) forms atleast 1 hairpin loop structures.
 24. The method of claim 22, wherein thetemplate RNA: a) further comprises a sequence comprising at least 20base pairs of at least 80% identity to a target DNA strand; b) furthercomprises a sequence comprising at least 20 base pairs of at least 80%identity to a target DNA strand is at the 5′ end of the template RNA c)has at least 3 bases of 100% identity to the target DNA strand.
 25. Themethod of claim 22, wherein the insert sequence is between 50-50,000base pairs.
 26. The method of claim 22, wherein the target DNA is agenomic safe harbor (GSH) site.
 27. A pharmaceutical composition,comprising the system of any of claim 1 and a pharmaceuticallyacceptable excipient or carrier, wherein the pharmaceutically acceptableexcipient or carrier is selected from a vector, a vesicle, and a lipidnanoparticle.